Language：English   Publishing type：Research paper (scientific journal)  

Background: Japanese cedar (Cryptomeria japonica) is an important tree for Japanese forestry. Male-sterile marker development in Japanese cedar would facilitate selection of male-sterile plus trees, addressing the widespread social problem of pollinosis and facilitating the identification of heterozygotes, which are useful for breeding. Results: This study used next-generation sequencing for single-nucleotide polymorphism discovery in libraries constructed from several organs, including male-sterile and male-fertile strobili. The single-nucleotide polymorphisms obtained were used to construct a high-density linkage map, which enabled identification of a locus on linkage group 9 strongly correlated with male-sterile trait. Expressed sequence tags corresponding to 11 marker loci from 5 isotigs were associated with this locus within 33.4-34.5 cM. These marker loci explained 100% of the phenotypic variation. Several homologs of these sequences are associated with male sterility in rice or Arabidopsis, including a pre-mRNA splicing factor, a DEAD-box protein, a glycosyl hydrolase, and a galactosyltransferase. These proteins are thus candidates for the causal male-sterile gene at the ms-1 locus. After we used a SNaPshot assay to develop markers for marker-assisted selection (MAS), we tested F₂ progeny between male-sterile and wild-type plus trees to validate the markers and extrapolated the testing to a larger plus-tree population. We found that two developed from one of the candidates for the causal gene were suitable for MAS. Conclusions: More than half of the ESTs and SNPs we collected were new, enlarging the genomic basis for genetic research on Japanese cedar. We developed two SNP markers aimed at MAS that distinguished individuals carrying the male-sterile trait with 100% accuracy, as well as individuals heterozygous at the male-sterile locus, even outside the mapping population. These markers should enable practical MAS for conifer breeding.

DOI： 10.1186/s12864-018-4581-5

Language：English   Publishing type：Research paper (scientific journal)  

Toxicodendron succedaneum: (L.) Kuntze is a tree cultivated for the production of sumac wax, which is extracted from the mesocarp. There are several hypotheses regarding the origin of T. succedaneum on mainland Japan. In this study, the geographical distribution of genetic variation in 13 wild populations on Honshu, Shikoku, Kyushu, and Ryukyu Islands, Japan was investigated and compared with that of individuals from continental Asia. Seven chloroplast DNA haplotypes of T. succedaneum were observed in Japan and could be divided into three lineages based on relatedness between haplotypes. One of these lineages was also observed in continental Asia, and the others were genetically distant from the haplotypes that originated on the continent, with one considered to have originated on the Ryukyu Islands, and the other on mainland Japan. The genetic diversity of both chloroplast and nuclear DNA was lower in populations from Ryukyu Islands than in populations from mainland Japan. Bayesian clustering based on nuclear genotypes showed a clear difference between the groups from Ryukyu Islands and mainland Japan. Based on approximate Bayesian computation analysis of polymorphic data for both genomes, it was inferred that wild populations of T. succedaneum on mainland Japan consist of both lineages with natural distribution on mainland Japan and those introduced from Ryukyu Islands and continental Asia.

DOI： 10.1007/s10265-017-0992-7

Language：English   Publishing type：Research paper (scientific journal)  

Although most molecular ecology studies examining genetic variation have focused on natural forests, for some major tree species, natural forests are nearly extinct, and the remaining genetic resources are mainly planted forests. In order to manage the genetic variability and develop a conservation strategy for such species, it is important to examine genetic variation resulting from historical processes during repeated artificial population development through plantations. The geographic pattern of genetic diversity and structure of 49 old planted Pinus thunbergii populations (2755 trees) distributed across Japan was examined using seven nuclear microsatellite markers. We found that allelic diversity was generally lower in both northern and eastern populations; however, locally, some populations in other regions also exhibited low allelic diversity. The overall value of the standardized measure of population differentiation (G′_ST = 0.206) was higher than that of both other widespread Japanese conifers and continental Pinus species. STRUCTURE software revealed a general gradual cline in genetic structure from southwestern to northeastern populations; however, some populations on the Japan Sea side showed quite a different local proportion of cluster memberships from nearby populations. These observations indicated that most of the preserved, planted populations of P. thunbergii possess regional genetic variation, but some populations were developed from seed pools derived from other regions, possibly through distribution by ship along the Japan Sea. Information on this specific genetic variation as a result of historical seed use and transfer should assist the design of several conservation units and breeding zones, while also taking care of a deep-seated need for conservation of pine forests by local people.

DOI： 10.1016/j.foreco.2017.10.026

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)  

DOI： 10.1093/treephys/tpw001

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)  

DOI： 10.1007/s11295-015-0859-9

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DOI： 10.1007/s10310-014-0460-3

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1186/s12870-014-0308-1

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1371/journal.pone.0079866

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Language：Others   Publishing type：Research paper (scientific journal)   Publisher：Journal of Forest Research  

To determine the potential reduction of the seed productivity of Japanese black pine (Pinus thunbergii) by the western conifer seed bug (Leptoglossus occidentalis), two feeding experiments were conducted in the center of Kyushu Island, southwestern Japan, in fall of 2020. First, L. occidentalis individuals and a preharvested P. thunbergii cone were enclosed in a nonwoven fabric bag for 2 weeks, which yielded a mean filled seed rate of 18.8% compared with that of 35.6% in the control. Second, L. occidentalis individuals and harvested cones were enclosed in a plastic case for 1 week, affording a mean filled seed rate of 25.7% compared with that of 36.4% in the control. These results suggest that L. occidentalis feeding reduces the filled seed rate of P. thunbergii and damages the seed orchards of this species. Additionally, P. thunbergii seeds from cones damaged by L. occidentalis could be distinguished by their germination ability when sorted using an ethanol floatation technique. Overall, these findings suggest the importance of carefully monitoring L. occidentalis infestation in P. thunbergii seed orchards.

DOI： 10.1080/13416979.2023.2291343

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Language：Japanese   Publisher：The Japanese Forest Society  

Simple Identifying Highly Productive Lacquer Tree (Toxicodendron vernicifluum) Clones by Measuring Resin Exudate Length, Tree Growth Traits, and Leaf Characteristics
Increased production and a stable domestic supply of tree lacquer from Toxicodendron vernicifluum is essential for restoring national treasures and important cultural properties in Japan. Therefore, it is beneficial to identify clones that produce large quantities of lacquer. In this study, we used DNA markers to investigate the clonal structures of lacquer trees originating from roots at seven sites in Ibaraki Prefecture. In addition, we measured the amount of lacquer produced by several tree within each clone group. We determined the relationships between resin exudate length, tree growth traits, and leaf characteristics to find indices that can be used to indirectly estimate the amounts of lacquer produced. In total, 10 clones (A-J) were detected at the sites, with about 50％ of individuals belonging to clone E. We found that individual trees with clone groups tended to share characteristics that differed among clones. In particular, the amounts of lacquer production and diameter at breast height (DBH) were significantly different between clones. Clones with larger DBH values produced larger amounts of lacquer. Even at different sites, the growth and leaf characteristics of individuals within each clone group were very similar. Our study suggested that measurement of exuded resin, DBH, and leaf characteristics in individuals around 10-20 years old can be used to identify clones that produce large amounts of lacquer.

DOI： 10.4005/jjfs.105.87

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Publishing type：Research paper (scientific journal)   Publisher：Ecological Research  

Forest trees species are often genetically adapted to local environmental conditions. Therefore, local seeds are recommended for ecological restoration. However, seedlings of broad-leaved tree species, such as Japanese beech (Fagus crenata), are limited in their commercial seedling production in Japan. Thus, long-distance transfer of seeds and/or seedlings is common. F. crenata's distinct geographical structure is well known; large-scale seed transfer may increase the risk of genetic disturbance. Several provenance trials of the species revealed that phenotypic traits such as leaf area and bud flush date differed latitudinally and/or between the Pacific Ocean side and the Japan Sea side. We investigated leaf size and bud flush date and identified the chloroplast DNA haplotype of trees planted in two provenance trials established in Hokkaido, Japan. We then examined whether cpDNA haplotype information is useful as a proxy in ecological restoration when using seedlings with incomplete seed source information. This study indicated that suitable seedlings could be selected based on chloroplast DNA haplotype information in F. crenata in the case of incomplete seed source information.

DOI： 10.1111/1440-1703.12351

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：PLoS ONE  

Environmental responses are critical for plant growth and survival under different climate conditions. To elucidate the underlying biological mechanisms of environmental responses in Japanese cedar (Cryptomeria japonica D. Don), the annual transcriptome dynamics of common clonal trees (Godai1) planted at three different climate sites (Yamagata, Ibaraki, and Kumamoto Prefectures) were analyzed using microarrays. Both principal component analysis (PCA) and hierarchical clustering of the microarray data indicated the transition to dormant transcriptome status occurred earlier and the transition to active growth status later in the colder region. Interestingly, PCA also indicated that the transcriptomes of trees grown under three different conditions were similar during the growth period (June to September), whereas the transcriptomes differed between sites during the dormant period (January to March). In between-site comparisons, analyses of the annual expression profiles of genes for sites 'Yamagata vs. Kumamoto', 'Yamagata vs. Ibaraki', and 'Ibaraki vs. Kumamoto' identified 1,473, 1,137, and 925 targets exhibiting significantly different expression patterns, respectively. The total of 2,505 targets that exhibited significantly different expression patterns in all three comparisons may play important roles in enabling cuttings to adapt to local environmental conditions. Partial least-squares regression analysis and Pearson correlation coefficient analysis revealed that air temperature and day length were the dominant factors controlling the expression levels of these targets. GO and Pfam enrichment analyses indicated that these targets include genes that may contribute to environmental adaptation, such as genes related to stress and abiotic stimulus responses. This study provided fundamental information regarding transcripts that may play an important role in adaptation to environmental conditions at different planting sites.

DOI： 10.1371/journal.pone.0277797

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Language：Others   Publishing type：Research paper (scientific journal)   Publisher：Tree Genetics and Genomes  

To achieve gene preservation in tree populations when planting seedlings to regenerate forests, specific and practical guidelines and criteria for seed collection are needed to ensure reliable coverage and effective capture of the current genetic variation. We examined the genetic variation of adult trees (524 trees) and seed pools (1618 seeds) collected from 70 mother trees in an old planted population of Pinus thunbergii in Japan using seven nuclear microsatellite markers. To consider a suitable seed collection strategy, we monitored changes in allelic diversity of seed pools with an increasing number of mother trees and examined the required number of mother trees. We found significantly higher allelic diversity statistics in seed pools of mother trees with smaller diameter at breast height. Spatial genetic autocorrelation analysis showed significantly positive kinship coefficients between both pairs of adult trees and seeds collected from mother trees for up to 100-m distance classes. Based on the rarefaction curve, seed pools obtained from approximately up to 30 mother trees could cover and saturate most of the genetic diversity and composition of the adult tree population and the potential overall seed pools, particularly for statistics that are less likely affected by rare alleles. Indications from the mother tree number, together with considerations for the size or spatial distribution of mother trees, are expected to contribute not only to guidelines for in situ genetic management of local planted populations, but also to the development of strategies for ex situ gene preservation of populations as genetic resources.

DOI： 10.1007/s11295-022-01584-5

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Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Others   Publishing type：Research paper (scientific journal)   Publisher：PLoS ONE  

The western conifer seed bug (WCSB, Leptoglossus occidentalis) is a pest of many pine species and is invasive worldwide. WCSB directly and indirectly deteriorates pine nut production by sucking seeds from cones. Currently, researchers think that WCSBs search for food by a combination of cues from visible light, infrared radiation, and chemicals such as monoterpenes. Some research revealed that WCSBs prefer larger cones, and it was thought that WCSBs suck seeds from and obtain more heat on larger cones. However, in early spring, we observed that most WCSBs gathered on male cones rather than on female cones and young cones. We hypothesized that male pine cones were warmer than female cones and needles, and WCSBs sucking male cones may receive more heat. To test these hypotheses, we measured spectral reflectance with a hyperspectral sensor and temperature of pine organs with tiny thermocouples, and the data were analyzed by a heat budget model. Our results revealed that male cones were significantly warmer and more reflective than female cones and needles, which may attract WCSBs. These results supported our hypothesis that WCSBs on male cones were warmer than those on other organs. This study will help further understanding of WCSBs and the adaptive value of pine cone colors.

DOI： 10.1371/journal.pone.0272565

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Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)   Publisher：THE JAPANESE FORESTRY SOCIETY  

Detection of Plus Trees Heterozygous for a Male-sterility Gene in Hinoki Cypress (Chamaecyparis obtusa (Sieb. et Zucc.) Endl.), Using Self-pollinated Families

DOI： 10.4005/jjfs.104.176

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Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Others   Publishing type：Research paper (scientific journal)  

Although cone morphology has been mainly utilized for taxonomy or forestry purposes, characterization of the environmental factors that influence its variation is still insufficient. Major conifers distributed over diverse climatic conditions like the Japanese archipelago could show cone morphology variation among populations related to large differences in key influential factors such as temperature, snow, irradiance and sunshine. Geographical variation in cone characteristics (cone size, seed production, seed productivity per cone and seed size) was examined across the species distribution in 24 old planted Pinus thunbergii populations (479 trees) along two major seasides in Japan (Pacific Ocean side and Japan Sea side). Variance components of cone characteristics explained by seaside were at similar levels to those for populations nested within seasides. Populations on the Japan Sea side produced cones that had larger size (length and width), higher seed production (number of filled seeds per cone and seed mass per cone) and seed productivity per cone (seed-to-ovule ratio). Analysis of covariance showed that significant latitudinal clines observed in cone size and seed productivity per cone were mostly a result of inter-seaside differences. Linear mixed model analysis detected that geographical variation in cone size, seed production and seed productivity per cone in the populations were affected significantly by low temperature, more snow, less solar irradiance and less sunshine time. Large cone sizes and high seed productivity per cone found in the populations on the Japan Sea side could be explained by a genetic or plastic response to maintain reproductive success.

DOI： 10.1111/1442-1984.12309

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1111/1442-1984.12309

Language：Others   Publishing type：Research paper (scientific journal)  

In the twentieth century, a substantial decline in Pinus thunbergii populations in Japan occurred due to the outbreak of pine wood nematode (PWN), Burshaphelencus xylophilus. A PWNP. thunbergii resistant trees-breeding project was developed in the 1980s to provide reforestation materials to minimalize the pest damage within the population. Since climate change can also contribute to PWN outbreaks, an intensive reforestation plan instated without much consideration can impact on the genetic diversity of P. thunbergii populations. The usage and deployment of PWN-P. thunbergii resistant trees to a given site without genetic management can lead to a genetic disturbance. The Iki-no-Matsubara population was used as a model to design an approach for the deployment management. This research aimed to preserve local genetic diversity, genetic structure, and relatedness by developing a method for deploying Kyushu PWN-P. thunbergii resistant trees as reforestation-material plants into Iki-no-Matsubara. The local genotypes of the Iki-no-Matsubara population and the Kyushu PWN-P. thunbergii resistant trees were analyzed using six microsatellite markers. Genotype origins, relatedness, diversity, and structure of both were investigated and compared with the genetic results previously obtained for old populations of P. thunbergii throughout Japan. A sufficient number of Kyushu PWN-P. thunbergii resistant trees, as mother trees, within seed orchards and sufficient status number of the seedlings to deploy are needed when deploying the Kyushu PWN-P. thunbergii resistant trees as reforestation material planting into Iki-no-Matsubara population. This approach not only be used to preserve Iki-no-Matsubara population (genetic diversity, genetic structure, relatedness, and resilience of the forests) but can also be applied to minimize PWN damage. These results provide a baseline for further seed sourcing as well as develop genetic management strategies within P. thunbergii populations, including Kyushu PWN-P. thunbergii resistant trees.

DOI： 10.3390/f12010072

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Others   Publishing type：Research paper (scientific journal)  

Pine wilt disease (PWD) caused by the pinewood nematode (PWN) (Bursaphelenchus xylophilus (Steiner and Buhrer) Nickle) is a worldwide issue. Infection is considered to be promoted mainly by the increased air temperature, but it is important to investigate whether the effect of high temperature similarly influences the different ranks of resistant clone. In the present study, we conducted PWN inoculation tests using six common open-pollinated families of resistant Pinus thunbergii Parl. The tests were conducted at nurseries of five test sites across Japanese archipelago between 2015 and 2017. Our analysis focused specifically on temperature. Firstly, we examined the effects of test sites, inoculation year, and their interaction on unaffected seedling rate and found that the unaffected seedling rate of all tested pine families decreased as the cumulative temperature increased. We found that the unaffected seedling rate decreased as the cumulative temperature increased for all tested pine families. In general, higher cumulative temperatures were required for having an effect on the unaffected seedling rates of higher PWN-resistant families. Typically, early cumulative temperatures, i.e., 19 days after inoculation, had the greatest effect on the unaffected seedling rates of PWN-resistant pines. However, the relationship between cumulative temperature and predicted unaffected seedling rate follow similar rate for all families. Thus, the order of resistance level is maintained in terms of the cumulative temperature required for having an effect.

DOI： 10.3390/F11090922

Language：Others   Publishing type：Research paper (scientific journal)  

BackgroundandObjectives:Todeterminewhether theprogenyofpinewoodnematode-resistant Pinus thunbergiiParl. clones selectedinthe southwesternregionof Japancouldbe successful inreforestation in the northern region, we investigated the magnitude of the genotype-environment interaction effect on the resistance against Bursaphelenchus xylophilus (Steiner and Buhrer) Nickle in P. thunbergii. Materials andMethods: We inoculated P. thunbergii seedlings of six full-sib families, with various resistance levels, with B. xylophilus in nurseries at three experimental sites in the northern and southern regions of Japan. All parental clones of the tested families originated from southwestern Japan, and selection of parental clones for resistance was performed in the same region. Sound rates after nematode inoculation were calculated, and survival analysis, correlation analysis and variance component analysis were performed. Results and Conclusions: Families with high sound rate in the southern region also showed a high sound rate in the northern region. In almost all cases, Spearman's correlation coefficients for sound rates were more than 0.698 among sites. The variance component of the interaction between site and family was small compared to that of site and family separately. Thus, we conclude that the resistant clones selected in the southern region would retain their genetic resistance in the northern regions.

DOI： 10.3390/f11090955

Language：English   Publishing type：Research paper (scientific journal)  

Language：Others   Publishing type：Research paper (scientific journal)  

In recent years, candidate regions containing causative genes of Sosyun, a male sterility trait, have been identified by a linkage map in Cryptomeria japonica with quantitative trait loci analysis. Of which, reCj19250 was found to be closely linked to the causative gene in this candidate region. Another gene called reCj11611, which is presumed to be related to carbohydrate metabolism, was mapped to the same locus as reCj19250, but no details about this gene have been reported to date. In this study, the gene expression of both reCj11611 and reCj19250, or the monosaccharide and starch content, were investigated in C. japonica clones during pollen formation. The monosaccharide or starch content during pollen formation increased significantly in Sosyun, suggesting that carbohydrate metabolism is abnormal in plants with this trait. However, evidence that gene expression of reCj11611 is related to abnormal carbohydrate metabolism was not obtained. A specific mutation of the reCj11611 cDNA sequence associated with Sosyun was determined. Deletion of 20 bps sequences in the 3′UTR region for the reCj11611 cDNA sequence was applied to approximately 760 C. japonica clones using a DNA marker which was able to identify this deletion. Although six clones were shown to be heterozygotic for this gene, five of them were incongruent with the presence of reCj19250. Therefore, the genes cited to be in the same position on the C. japonica linkage map were, in fact, at neighboring loci.

DOI： 10.4005/jjfs.102.191

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Others   Publishing type：Research paper (scientific journal)  

Seasonal phenomena in plants are primarily affected by day length and temperature. The shoot transcriptomes of trees grown in the field and a controlled-environment chamber were compared to characterize genes that control annual rhythms and the effects of day length- and temperature-regulated genes in the gymnosperm Japanese cedar (Cryptomeria japonica D. Don), which exhibits seasonally indeterminate growth. Annual transcriptome dynamics were clearly demonstrated by principal component analysis using microarray data obtained under field-grown conditions. Analysis of microarray data from trees grown in a controlled chamber identified 2,314 targets exhibiting significantly different expression patterns under short-day (SD) and long-day conditions, and 2,045 targets exhibited significantly different expression patterns at 15°C (LT; low temperature) versus 25°C. Interestingly, although growth was suppressed under both SD and LT conditions, approximately 80% of the SD- and LT-regulated targets differed, suggesting that each factor plays a unique role in the annual cycle. The top 1,000 up-regulated targets in the growth/dormant period in the field coincided with more than 50% of the SD- and LT-regulated targets, and gene co-expression network analysis of the annual transcriptome indicated a close relationship between the SD- and LT-regulated targets. These results indicate that the respective effects of day length and temperature interact to control annual transcriptome dynamics. Well-known upstream genes of signaling pathways responsive to environmental conditions, such as the core clock (LHY/CjLHYb and CCA1/CjLHYa) and PEBP family (MFT) genes, exhibited unique expression patterns in Japanese cedar compared with previous reports in other species, suggesting that these genes control differences in seasonal regulation mechanisms between species. The results of this study provide new insights into seasonal regulation of transcription in Japanese cedar.

DOI： 10.1371/journal.pone.0229843

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Pine wilt disease (PWD) affects forests globally and is caused by the pinewood nematode (PWN) Bursaphelenchus xylophilus. It has been proposed that PWD development has a strong association with air temperature. Because symptom development is related to the number of PWNs and they are poikilothermic animals whose development and multiplication are affected by temperature; it appears that temperature conditions affect the PWN multiplication rate, which leads to difference in disease progression. However, limited information is available about the temperatures inside pine trees, and the understanding of how temperature affects PWN multiplication and PWD progression remains incomplete. Therefore, in the present study, we evaluated the temperature, number of PWNs, and external symptoms of PWD in Pinus thunbergii seedlings infected with PWNs at air temperatures of 20 °C, 25 °C, and 30 °C over time. We found that the seedling temperatures were slightly higher than the air temperature under each temperature condition and that the effective accumulated temperature calculated using a base temperature of 10 °C was related to the number of PWNs and symptom development in all temperature test groups. However, at 20 °C, there were very few PWNs in some seedlings and none of the seedlings showed partial or full wilting. These findings suggest that PWD progression is greatly affected by ambient temperature and facilitated by increase in PWN populations with effective accumulated temperature especially in the range of 25 °C–30 °C.

DOI： 10.1007/s10658-019-01908-7

Language：Others   Publishing type：Research paper (scientific journal)  

Pine wilt disease (PWD) affects forests globally and is caused by the pinewood nematode (PWN) Bursaphelenchus xylophilus. It has been proposed that PWD development has a strong association with air temperature. Because symptom development is related to the number of PWNs and they are poikilothermic animals whose development and multiplication are affected by temperature; it appears that temperature conditions affect the PWN multiplication rate, which leads to difference in disease progression. However, limited information is available about the temperatures inside pine trees, and the understanding of how temperature affects PWN multiplication and PWD progression remains incomplete. Therefore, in the present study, we evaluated the temperature, number of PWNs, and external symptoms of PWD in Pinus thunbergii seedlings infected with PWNs at air temperatures of 20 °C, 25 °C, and 30 °C over time. We found that the seedling temperatures were slightly higher than the air temperature under each temperature condition and that the effective accumulated temperature calculated using a base temperature of 10 °C was related to the number of PWNs and symptom development in all temperature test groups. However, at 20 °C, there were very few PWNs in some seedlings and none of the seedlings showed partial or full wilting. These findings suggest that PWD progression is greatly affected by ambient temperature and facilitated by increase in PWN populations with effective accumulated temperature especially in the range of 25 °C–30 °C.

DOI： 10.1007/s10658-019-01908-7

Language：English   Publishing type：Research paper (scientific journal)  

Seasonal phenomena in plants are primarily affected by day length and temperature. The shoot transcriptomes of trees grown in the field and a controlled-environment chamber were compared to characterize genes that control annual rhythms and the effects of day length- and temperature-regulated genes in the gymnosperm Japanese cedar (Cryptomeria japonica D. Don), which exhibits seasonally indeterminate growth. Annual transcriptome dynamics were clearly demonstrated by principal component analysis using microarray data obtained under field-grown conditions. Analysis of microarray data from trees grown in a controlled chamber identified 2,314 targets exhibiting significantly different expression patterns under short-day (SD) and long-day conditions, and 2,045 targets exhibited significantly different expression patterns at 15°C (LT; low temperature) versus 25°C. Interestingly, although growth was suppressed under both SD and LT conditions, approximately 80% of the SD- and LT-regulated targets differed, suggesting that each factor plays a unique role in the annual cycle. The top 1,000 up-regulated targets in the growth/dormant period in the field coincided with more than 50% of the SD- and LT-regulated targets, and gene co-expression network analysis of the annual transcriptome indicated a close relationship between the SD- and LT-regulated targets. These results indicate that the respective effects of day length and temperature interact to control annual transcriptome dynamics. Well-known upstream genes of signaling pathways responsive to environmental conditions, such as the core clock (LHY/CjLHYb and CCA1/CjLHYa) and PEBP family (MFT) genes, exhibited unique expression patterns in Japanese cedar compared with previous reports in other species, suggesting that these genes control differences in seasonal regulation mechanisms between species. The results of this study provide new insights into seasonal regulation of transcription in Japanese cedar.

DOI： 10.1371/journal.pone.0229843

Language：Japanese   Publishing type：Research paper (scientific journal)  

DOI： https://doi.org/10.4005/jjfs.101.298

Language：English   Publishing type：Research paper (scientific journal)  

Background: Pine wilt disease (PWD), which is caused by the pine wood nematode (PWN) Bursaphelenchus xylophilus, is currently the greatest threat to pine forests in Europe and East Asian countries including Japan. Constructing a detailed linkage map of DNA markers and identifying PWD resistance genes/loci lead to improved resistance in Pinus thunbergii, as well as other Pinus species that are also susceptible to PWD. Results: A total F₁ mapping population of 188 individuals derived from a cross between the PWD-resistant P. thunbergii varieties 'Tanabe 54' (resistant rank 2 to PWD) and 'Tosashimizu 63' (resistant rank 4 to PWD) was inoculated with PWN, and was evaluated for disease symptoms. To perform linkage analysis for PWN resistance, a set of three maps was constructed; two parental maps generated using the integrated two-way pseudo-testcross method, and a consensus map with population-type cross-pollination. The linkage map of 'Tanabe 54' consisted of 167 loci, and covered 14 linkage groups (LGs), with a total genetic distance of 1214.6 cM. The linkage map of 'Tosashimizu 63' consisted of 252 loci, and covered 14 LGs, with a total genetic distance of 1422.1 cM. The integrated consensus map comprised 12 LGs with the basic chromosome number of P. thunbergii, and a total genetic distance of 1403.6 cM. Results from quantitative trait loci (QTL) analysis using phenotype data and linkage maps indicated that PWN resistance is controlled by a single dominant allele, which was derived from the 'Tanabe 54' female parent. This major QTL was located on linkage group 3 and was designated PWD1 for PINE WILT DISEASE 1. Conclusions: The PWD1 locus is a major resistance QTL located on the Pinus consensus LG03 that acts in a dominant manner to confer pine wood nematode resistance. Information from the present study will be useful for P. thunbergii breeding programs to improve resistance to PWD, and also to help identify susceptibility genes in Pinus species.

DOI： 10.1186/s12870-019-2045-y

Language：Others   Publishing type：Research paper (scientific journal)  

Background: Pine wilt disease (PWD), which is caused by the pine wood nematode (PWN) Bursaphelenchus xylophilus, is currently the greatest threat to pine forests in Europe and East Asian countries including Japan. Constructing a detailed linkage map of DNA markers and identifying PWD resistance genes/loci lead to improved resistance in Pinus thunbergii, as well as other Pinus species that are also susceptible to PWD. Results: A total F1 mapping population of 188 individuals derived from a cross between the PWD-resistant P. thunbergii varieties 'Tanabe 54' (resistant rank 2 to PWD) and 'Tosashimizu 63' (resistant rank 4 to PWD) was inoculated with PWN, and was evaluated for disease symptoms. To perform linkage analysis for PWN resistance, a set of three maps was constructed; two parental maps generated using the integrated two-way pseudo-testcross method, and a consensus map with population-type cross-pollination. The linkage map of 'Tanabe 54' consisted of 167 loci, and covered 14 linkage groups (LGs), with a total genetic distance of 1214.6 cM. The linkage map of 'Tosashimizu 63' consisted of 252 loci, and covered 14 LGs, with a total genetic distance of 1422.1 cM. The integrated consensus map comprised 12 LGs with the basic chromosome number of P. thunbergii, and a total genetic distance of 1403.6 cM. Results from quantitative trait loci (QTL) analysis using phenotype data and linkage maps indicated that PWN resistance is controlled by a single dominant allele, which was derived from the 'Tanabe 54' female parent. This major QTL was located on linkage group 3 and was designated PWD1 for PINE WILT DISEASE 1. Conclusions: The PWD1 locus is a major resistance QTL located on the Pinus consensus LG03 that acts in a dominant manner to confer pine wood nematode resistance. Information from the present study will be useful for P. thunbergii breeding programs to improve resistance to PWD, and also to help identify susceptibility genes in Pinus species.

DOI： 10.1186/s12870-019-2045-y

Language：Others   Publishing type：Research paper (scientific journal)  

Pine wilt disease is of major concern as it has destroyed pine forests in East Asia and Europe. Several studies have suggested that invasion by the pinewood nematode (PWN) Bursaphelenchus xylophilus, which causes this disease, evokes an excessive defence response in pine trees, resulting in tree death. However, few studies have quantitatively evaluated the correlation between PWN distribution and tree defence responses. Therefore, the present study aimed to quantify the number of PWNs and expression levels of putative pathogenesis-related (PR) genes in different positions of Japanese black pine (Pinus thunbergii) seedlings over time. To quantify the number of PWNs in the seedlings, we used TaqMan quantitative real-time PCR (qPCR) assay. During the early phase of infection, most PWNs were distributed around the inoculated sites, with only a small number being detected at distant sites, but the expression levels of PR genes were highly upregulated throughout the seedlings. Both the number of PWNs and expression levels of PR genes then increased drastically throughout the seedlings, all of which exhibited external symptoms. Thus, it appears that the rapid migration of PWNs induces a defence response throughout the seedling; however, this may not be effective in controlling these parasites, thereby ultimately leading to plant death.

DOI： 10.1111/efp.12518

Language：Japanese   Publishing type：Research paper (scientific journal)  

Development of simple DNA markers for selecting trees with the male-sterile gene of Cryptomeria japonica “Sosyun”
We report the development of two types of simple DNA markers that can be used to easily identify genotypes of both the male-sterile and wild-type individuals at a locus, which is tightly linked to the causative gene of male sterility in the “Sosyun” clone of Cryptomeria japonica. Both the allele-specific primer (ASP) PCR marker and cleaved amplified polymorphic sequence (CAPS) marker were designed using previously reported single nucleotide polymorphism (SNP) information. The cDNA and genomic DNA of the EST reCj19250, which is linked to the ms-1 locus, were sequenced for both the “Sosyun” and wild-type plus-tree “Usui2” clones. Primers were designed for detecting both the sterile and wild-type SNPs using the genomic DNA sequences. For the ASP-PCR marker, homozygous recessive male-sterile individuals yielded a single 157-bp fragment, whereas the homozygous wild-type individuals yielded a single 345-bp fragment, and heterozygotes yielded both fragments. Similarly, for the CAPS marker, male-sterile individuals yielded a single 101-bp fragment after digestion, whereas wild-type ones yielded an intact single 125-bp fragment, and heterozygotes yielded fragments of both sizes. These markers could be utilized for identifying male-sterile individuals during seedling production and for identifying heterozygous trees for future tree breeding.

DOI： 10.4005/jjfs.101.155

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

We report the development of two types of simple DNA markers that can be used to easily identify genotypes of both the male-sterile and wild-type individuals at a locus, which is tightly linked to the causative gene of male sterility in the “Sosyun” clone of Cryptomeria japonica. Both the allele-specific primer (ASP) PCR marker and cleaved amplified polymorphic sequence (CAPS) marker were designed using previously reported single nucleotide polymorphism (SNP) information. The cDNA and genomic DNA of the EST reCj 19250, which is linked to the ms-1 locus, were sequenced for both the “Sosyun” and wild-type plus-tree “Usui 2” clones. Primers were designed for detecting both the sterile and wild-type SNPs using the genomic DNA sequences. For the ASP-PCR marker, homozygous recessive male-sterile individuals yielded a single 157-bp fragment, whereas the homozygous wild-type individuals yielded a single 345-bp fragment, and heterozygotes yielded both fragments. Similarly, for the CAPS marker, male-sterile individuals yielded a single 101-bp fragment after digestion, whereas wild-type ones yielded an intact single 125 -bp fragment, and heterozygotes yielded fragments of both sizes. These markers could be utilized for identifying male-sterile individuals during seedling production and for identifying heterozygous trees for future tree breeding.

DOI： 10.4005/jjfs.101.155

Language：Japanese   Publishing type：Research paper (scientific journal)  

DOI： https://doi.org/10.4005/jjfs.100.218

Language：English   Publishing type：Research paper (scientific journal)  

The transport of monolignols from the cytosol to the cell wall is essential for lignin synthesis. The ATP-binding cassette (ABC) transporters may be involved in the transport of lignin precursors. ABC transporter genes subjected to expression analysis were chosen based on two criteria for screening candidate transporter genes related to lignification. The expression levels of 15 target genes in five plant organs were analyzed by real-time PCR. Five transporter genes (ABCG29, ABCG30, ABCG33, ABCG34, and ABCG37), which were simultaneously expressed with the reference genes, were selected as candidates. The candidate gene expression levels in root tissues of T-DNA insertion mutants were determined by semi-quantitative reverse transcription PCR. ABCG30 was more highly expressed in the abcg34 mutant than in the wild-type plants, while the expression of ABCG34 was twofold higher in the abcg30 mutant plants than in the wild-type plants. Thus, the expression of ABCG30 and ABCG34 may affect each other. There was no significant change in lignin content and composition in the single-gene knockout mutants of the candidate transporter genes, which suggested that each candidate gene did not solely contribute to lignin synthesis.

DOI： 10.1007/s10086-018-1733-9

Language：Others   Publishing type：Research paper (scientific journal)  

The transport of monolignols from the cytosol to the cell wall is essential for lignin synthesis. The ATP-binding cassette (ABC) transporters may be involved in the transport of lignin precursors. ABC transporter genes subjected to expression analysis were chosen based on two criteria for screening candidate transporter genes related to lignification. The expression levels of 15 target genes in five plant organs were analyzed by real-time PCR. Five transporter genes (ABCG29, ABCG30, ABCG33, ABCG34, and ABCG37), which were simultaneously expressed with the reference genes, were selected as candidates. The candidate gene expression levels in root tissues of T-DNA insertion mutants were determined by semi-quantitative reverse transcription PCR. ABCG30 was more highly expressed in the abcg34 mutant than in the wild-type plants, while the expression of ABCG34 was twofold higher in the abcg30 mutant plants than in the wild-type plants. Thus, the expression of ABCG30 and ABCG34 may affect each other. There was no significant change in lignin content and composition in the single-gene knockout mutants of the candidate transporter genes, which suggested that each candidate gene did not solely contribute to lignin synthesis.

DOI： 10.1007/s10086-018-1733-9

Language：English   Publishing type：Research paper (scientific journal)  

A genome-wide association study (GWAS) was conducted on more than 30,000 single nucleotide polymorphisms (SNPs) in unrelated first-generation plus tree genotypes from three populations of Japanese cedar Cryptomeria japonica D. Don with genomic prediction for traits of growth, wood properties and male fecundity. Among the assessed populations, genetic characteristics including the extent of linkage disequilibrium (LD) and genetic structure differed and these differences are considered to be due to differences in genetic background. Through population-independent GWAS, several significant SNPs found close to the regions associated with each of these traits and shared in common across the populations were identified. The accuracies of genomic predictions were dependent on the traits and populations and reflected the genetic architecture of traits and genetic characteristics. Prediction accuracies using SNPs selected based on GWAS results were similar to those using all SNPs for several combinations of traits and populations. We discussed the application of genome-wide studies for C. japonica improvement.

DOI： 10.3389/fpls.2018.01322

Language：Others   Publishing type：Research paper (scientific journal)  

Background: Adventitious root formation is an essential physiological process for successful propagation of cuttings in various plant species. Because coniferous species are highly heterozygous, propagation of cuttings is of great practical use in breeding. Although various factors influence adventitious root formation, little is known of the associated regulatory mechanisms. Whereas adventitious roots generally form from the base of cuttings, this process is accompanied by physiological changes in leaves, which supply assimilates and metabolites. Herein, we present microarray analyses of transcriptome dynamics during adventitious root formation in whole cuttings in the coniferous species, Cryptomeria japonica. Results: Temporal patterns of gene expression were determined in the base, the middle, and needles of cuttings at eight time points during adventitious root formation. Global gene expression at the base had diverged from that in the middle by 3-h post-insertion, and changed little in the subsequent 3-days post-insertion, and global gene expression in needles altered characteristically at 3- and 6-weeks post-insertion. In Gene Ontology enrichment analysis of major gene clusters based on hierarchical clustering, the expression profiles of genes related to carbohydrates, plant hormones, and other categories indicated multiple biological changes that were involved in adventitious root formation. Conclusions: The present comprehensive transcriptome analyses indicate major transcriptional turning and contribute to the understanding of the biological processes and molecular factors that influence adventitious root formation in C. japonica.

DOI： 10.1186/s12870-018-1401-7

Language：Others   Publishing type：Research paper (scientific journal)  

A genome-wide association study (GWAS) was conducted on more than 30,000 single nucleotide polymorphisms (SNPs) in unrelated first-generation plus tree genotypes from three populations of Japanese cedar Cryptomeria japonica D. Don with genomic prediction for traits of growth, wood properties and male fecundity. Among the assessed populations, genetic characteristics including the extent of linkage disequilibrium (LD) and genetic structure differed and these differences are considered to be due to differences in genetic background. Through population-independent GWAS, several significant SNPs found close to the regions associated with each of these traits and shared in common across the populations were identified. The accuracies of genomic predictions were dependent on the traits and populations and reflected the genetic architecture of traits and genetic characteristics. Prediction accuracies using SNPs selected based on GWAS results were similar to those using all SNPs for several combinations of traits and populations. We discussed the application of genome-wide studies for C. japonica improvement.

DOI： 10.3389/fpls.2018.01322

Language：English   Publishing type：Research paper (scientific journal)  

Background: Adventitious root formation is an essential physiological process for successful propagation of cuttings in various plant species. Because coniferous species are highly heterozygous, propagation of cuttings is of great practical use in breeding. Although various factors influence adventitious root formation, little is known of the associated regulatory mechanisms. Whereas adventitious roots generally form from the base of cuttings, this process is accompanied by physiological changes in leaves, which supply assimilates and metabolites. Herein, we present microarray analyses of transcriptome dynamics during adventitious root formation in whole cuttings in the coniferous species, Cryptomeria japonica. Results: Temporal patterns of gene expression were determined in the base, the middle, and needles of cuttings at eight time points during adventitious root formation. Global gene expression at the base had diverged from that in the middle by 3-h post-insertion, and changed little in the subsequent 3-days post-insertion, and global gene expression in needles altered characteristically at 3- and 6-weeks post-insertion. In Gene Ontology enrichment analysis of major gene clusters based on hierarchical clustering, the expression profiles of genes related to carbohydrates, plant hormones, and other categories indicated multiple biological changes that were involved in adventitious root formation. Conclusions: The present comprehensive transcriptome analyses indicate major transcriptional turning and contribute to the understanding of the biological processes and molecular factors that influence adventitious root formation in C. japonica.

DOI： 10.1186/s12870-018-1401-7

Language：Others   Publishing type：Research paper (scientific journal)  

Background: Japanese cedar (Cryptomeria japonica) is an important tree for Japanese forestry. Male-sterile marker development in Japanese cedar would facilitate selection of male-sterile plus trees, addressing the widespread social problem of pollinosis and facilitating the identification of heterozygotes, which are useful for breeding. Results: This study used next-generation sequencing for single-nucleotide polymorphism discovery in libraries constructed from several organs, including male-sterile and male-fertile strobili. The single-nucleotide polymorphisms obtained were used to construct a high-density linkage map, which enabled identification of a locus on linkage group 9 strongly correlated with male-sterile trait. Expressed sequence tags corresponding to 11 marker loci from 5 isotigs were associated with this locus within 33.4-34.5 cM. These marker loci explained 100% of the phenotypic variation. Several homologs of these sequences are associated with male sterility in rice or Arabidopsis, including a pre-mRNA splicing factor, a DEAD-box protein, a glycosyl hydrolase, and a galactosyltransferase. These proteins are thus candidates for the causal male-sterile gene at the ms-1 locus. After we used a SNaPshot assay to develop markers for marker-assisted selection (MAS), we tested F2 progeny between male-sterile and wild-type plus trees to validate the markers and extrapolated the testing to a larger plus-tree population. We found that two developed from one of the candidates for the causal gene were suitable for MAS. Conclusions: More than half of the ESTs and SNPs we collected were new, enlarging the genomic basis for genetic research on Japanese cedar. We developed two SNP markers aimed at MAS that distinguished individuals carrying the male-sterile trait with 100% accuracy, as well as individuals heterozygous at the male-sterile locus, even outside the mapping population. These markers should enable practical MAS for conifer breeding.

DOI： 10.1186/s12864-018-4581-5

Language：English   Publishing type：Research paper (scientific journal)  

The mechanism of monolignol transportation from the cytosol to the apoplast is still unclear despite being an essential step of lignification. Recently, ATP-binding cassette (ABC) transporters were suggested to be involved in monolignol transport. However, there are no reliable clues to the transporters of the major lignin monomers coniferyl and synapyl alcohol. In this study, the lignification progress of Arabidopsis cultured cells during tracheary element differentiation was monitored. The expression of selected transporter genes, as well as lignification and cell-wall formation related genes as references, in differentiating cultured cell samples harvested at 2-day intervals was analyzed by real-time PCR and the data were statistically processed. The cell wall formation transcription factor MYB46, programmed-cell death related gene XCP1 and lignin polymerization peroxidase AtPrx25 were classified into the same cluster. Furthermore, the cluster closest to the abovementioned cluster contained the lignin synthesis transcription factor MYB58 and the Arabidopsis ABC transporters ABCG11, ABCG22, ABCG36 and ABCG29. This result suggested that these four ABC transporters may be involved in lignification. In the expression analysis, unexpectedly, the lignification-related genes CAD5 and C4H were not included in the same cluster as MYB58 and AtPrx25. The expression data also suggested that the lignification of tracheary elements in the culture, where lignification ratio finally reached to around 40%, continued after cell death because lignification actively progressed after programmed cell death-related gene started to be expressed.

DOI： 10.1007/s10265-017-0979-4

Language：Others   Publishing type：Research paper (scientific journal)  

Toxicodendron succedaneum: (L.) Kuntze is a tree cultivated for the production of sumac wax, which is extracted from the mesocarp. There are several hypotheses regarding the origin of T. succedaneum on mainland Japan. In this study, the geographical distribution of genetic variation in 13 wild populations on Honshu, Shikoku, Kyushu, and Ryukyu Islands, Japan was investigated and compared with that of individuals from continental Asia. Seven chloroplast DNA haplotypes of T. succedaneum were observed in Japan and could be divided into three lineages based on relatedness between haplotypes. One of these lineages was also observed in continental Asia, and the others were genetically distant from the haplotypes that originated on the continent, with one considered to have originated on the Ryukyu Islands, and the other on mainland Japan. The genetic diversity of both chloroplast and nuclear DNA was lower in populations from Ryukyu Islands than in populations from mainland Japan. Bayesian clustering based on nuclear genotypes showed a clear difference between the groups from Ryukyu Islands and mainland Japan. Based on approximate Bayesian computation analysis of polymorphic data for both genomes, it was inferred that wild populations of T. succedaneum on mainland Japan consist of both lineages with natural distribution on mainland Japan and those introduced from Ryukyu Islands and continental Asia.

DOI： 10.1007/s10265-017-0992-7

Language：Others   Publishing type：Research paper (scientific journal)  

The mechanism of monolignol transportation from the cytosol to the apoplast is still unclear despite being an essential step of lignification. Recently, ATP-binding cassette (ABC) transporters were suggested to be involved in monolignol transport. However, there are no reliable clues to the transporters of the major lignin monomers coniferyl and synapyl alcohol. In this study, the lignification progress of Arabidopsis cultured cells during tracheary element differentiation was monitored. The expression of selected transporter genes, as well as lignification and cell-wall formation related genes as references, in differentiating cultured cell samples harvested at 2-day intervals was analyzed by real-time PCR and the data were statistically processed. The cell wall formation transcription factor MYB46, programmed-cell death related gene XCP1 and lignin polymerization peroxidase AtPrx25 were classified into the same cluster. Furthermore, the cluster closest to the abovementioned cluster contained the lignin synthesis transcription factor MYB58 and the Arabidopsis ABC transporters ABCG11, ABCG22, ABCG36 and ABCG29. This result suggested that these four ABC transporters may be involved in lignification. In the expression analysis, unexpectedly, the lignification-related genes CAD5 and C4H were not included in the same cluster as MYB58 and AtPrx25. The expression data also suggested that the lignification of tracheary elements in the culture, where lignification ratio finally reached to around 40%, continued after cell death because lignification actively progressed after programmed cell death-related gene started to be expressed.

DOI： 10.1007/s10265-017-0979-4

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Others   Publishing type：Research paper (scientific journal)  

Although most molecular ecology studies examining genetic variation have focused on natural forests, for some major tree species, natural forests are nearly extinct, and the remaining genetic resources are mainly planted forests. In order to manage the genetic variability and develop a conservation strategy for such species, it is important to examine genetic variation resulting from historical processes during repeated artificial population development through plantations. The geographic pattern of genetic diversity and structure of 49 old planted Pinus thunbergii populations (2755 trees) distributed across Japan was examined using seven nuclear microsatellite markers. We found that allelic diversity was generally lower in both northern and eastern populations; however, locally, some populations in other regions also exhibited low allelic diversity. The overall value of the standardized measure of population differentiation (G′ST = 0.206) was higher than that of both other widespread Japanese conifers and continental Pinus species. STRUCTURE software revealed a general gradual cline in genetic structure from southwestern to northeastern populations; however, some populations on the Japan Sea side showed quite a different local proportion of cluster memberships from nearby populations. These observations indicated that most of the preserved, planted populations of P. thunbergii possess regional genetic variation, but some populations were developed from seed pools derived from other regions, possibly through distribution by ship along the Japan Sea. Information on this specific genetic variation as a result of historical seed use and transfer should assist the design of several conservation units and breeding zones, while also taking care of a deep-seated need for conservation of pine forests by local people.

DOI： 10.1016/j.foreco.2017.10.026

Language：English   Publishing type：Research paper (scientific journal)  

In order to predict the effects of future atmospheric conditions on forest productivity, it is necessary to clarify the physiological responses of major forest tree species to high concentrations of ozone (O 3) and carbon dioxide (CO 2). Furthermore, intraspecific variation of these responses should also be examined in order to predict productivity gains through tree improvements in the future. We investigated intraspecific variation in growth and photosynthesis of Cryptomeria japonica D. Don, a major silviculture species in Japan, in response to elevated concentrations of O 3 (eO 3) and CO 2 (eCO 2), separately and in combination. Cuttings of C. japonica were grown and exposed to two levels of O 3 (ambient and twice-ambient levels) in combination with two levels of CO 2 (ambient and 550 μmol mol '1 in the daytime) for two growing seasons in a free-air CO 2 enrichment experiment. There was no obvious negative effect of eO 3 on growth or photosynthetic traits of the C. japonica clones, but a positive effect was observed for annual height increments in the first growing season. Dry mass production and the photosynthetic rate increased under eCO 2 conditions, while the maximum carboxylation rate decreased. Significant interaction effects of eO 3 and eCO 2 on growth and photosynthetic traits were not observed. Clonal effects on growth and photosynthetic traits were significant, but the interactions between clones and O 3 and/or CO 2 treatments were not. Spearman's rank correlation coefficients between growth traits under ambient conditions and for each treatment were significantly positive, implying that clonal ranking in growth abilities might not be affected by either eO 3 or eCO 2. The knowledge obtained from this study will be helpful for species selection in afforestation programs, to continue and to improve current programs involving this species, and to accurately predict the CO 2 fixation capacity of Japanese forests.

DOI： 10.1093/treephys/tpx028

Language：Others   Publishing type：Research paper (scientific journal)  

In order to predict the effects of future atmospheric conditions on forest productivity, it is necessary to clarify the physiological responses of major forest tree species to high concentrations of ozone (O 3) and carbon dioxide (CO 2). Furthermore, intraspecific variation of these responses should also be examined in order to predict productivity gains through tree improvements in the future. We investigated intraspecific variation in growth and photosynthesis of Cryptomeria japonica D. Don, a major silviculture species in Japan, in response to elevated concentrations of O 3 (eO 3) and CO 2 (eCO 2), separately and in combination. Cuttings of C. japonica were grown and exposed to two levels of O 3 (ambient and twice-ambient levels) in combination with two levels of CO 2 (ambient and 550 μmol mol '1 in the daytime) for two growing seasons in a free-air CO 2 enrichment experiment. There was no obvious negative effect of eO 3 on growth or photosynthetic traits of the C. japonica clones, but a positive effect was observed for annual height increments in the first growing season. Dry mass production and the photosynthetic rate increased under eCO 2 conditions, while the maximum carboxylation rate decreased. Significant interaction effects of eO 3 and eCO 2 on growth and photosynthetic traits were not observed. Clonal effects on growth and photosynthetic traits were significant, but the interactions between clones and O 3 and/or CO 2 treatments were not. Spearman's rank correlation coefficients between growth traits under ambient conditions and for each treatment were significantly positive, implying that clonal ranking in growth abilities might not be affected by either eO 3 or eCO 2. The knowledge obtained from this study will be helpful for species selection in afforestation programs, to continue and to improve current programs involving this species, and to accurately predict the CO 2 fixation capacity of Japanese forests.

DOI： 10.1093/treephys/tpx028

Language：Others   Publishing type：Research paper (scientific journal)  

In the original publication of this article [1], the accession number was listed as DC882454 through DC883482 on page 27; instead, the number should be DRA000525.

DOI： 10.1186/s12864-016-3156-6

Language：English   Publishing type：Research paper (scientific journal)  

Language：Others   Publishing type：Research paper (scientific journal)  

Cryptomeria japonica is one of the most important forest tree species in Japan. To increase the demand for domestic timber, broad uses related to the functional attributes of C. japonica must be developed. Several studies have examined the usefulness of C. japonica in terms of its biological activities, but a comprehensive study subjecting all parts of the C. japonica plant to the same solvent for extraction has not been done. Here, methanol extractions from the leaves, branches, bearing branches, male flowers, female flowers, cones, bark, heartwood, sapwood, pith, rhizomes, roots, and pollen were subjected to several in vitro assays of their biological activities such as antioxidant activity, anti-lipase activity, antibacterial activity and melanin-biosynthesis-inhibition activity. Their total phenolic content was also determined. The methanol extracts from each part of C. japonica except for pollen showed strong activities in the bioactivity assays. Furthermore, the methanol extracts were analyzed by GC/MS. The phytochemical profile varied among extracts from various parts of C. japonica. Our results suggest the great potential of C. japonica for use as a functional ingredient in health-related products.

DOI： 10.1177/1934578x1601100939

Language：English   Publishing type：Research paper (scientific journal)  

We analyzed the effect of genotype and planting density on the growth patterns of height and diameter at breast height (DBH) and the efficiency of indirect early selection targeting on the low planting density using stem analysis data obtained from a test site composed of 6 cultivars and 3 planting densities (5, 000, 3, 000, 1, 500 trees/ha) at stand age 28. The growth pattern of height was mainly affected by genotype, but the effect of planting density was small. The growth pattern of DBH was affected by both genotype and planting density. Our results showed the importance of selection of the genotype of seedlings for the control of growth in plantations of Sugi. The results of genetic evaluation obtained at general planting density (3, 000 trees/ha) could be applied for the prediction of genetic performance of growth traits at different planting densities. Approximately 15 years would be required to obtain 80% of genetic gain at stand age 28 with the plantation of 1, 500 trees/ha according to the test results obtained at the plantation of general planting density (3, 000 trees/ha).

DOI： https://doi.org/10.4005/jjfs.98.45

Language：Others   Publishing type：Research paper (scientific journal)  

The molecular mechanisms that control male strobilus development in conifers are largely unknown because the developmental stages and related genes have not yet been characterized. The determination of male strobilus developmental stages will contribute to genetic research and reproductive biology in conifers. Our objectives in this study were to determine the developmental stages of male strobili by cytological and transcriptome analysis, and to determine the stages at which aberrant morphology is observed in a male-sterile mutant of Cryptomeria japonica D. Don to better understand the molecular mechanisms that control male strobilus and pollen development. Male strobilus development was observed for 8 months, from initiation to pollen dispersal. A set of 19,209 expressed sequence tags (ESTs) collected from a male reproductive library and a pollen library was used for microarray analysis. We divided male strobilus development into 10 stages by cytological and transcriptome analysis. Eight clusters (7324 ESTs) exhibited major changes in transcriptome profiles during male strobili and pollen development in C. japonica. Two clusters showed a gradual increase and decline in transcript abundance, respectively, while the other six clusters exhibited stage-specific changes. The stages at which the male sterility trait of Sosyun was expressed were identified using information on male strobilus and pollen developmental stages and gene expression profiles. Aberrant morphology was observed cytologically at Stage 6 (microspore stage), and differences in expression patterns compared with wild type were observed at Stage 4 (tetrad stage).

DOI： 10.1093/treephys/tpw001

Language：Japanese   Publishing type：Research paper (scientific journal)  

The effect of genotype and planting density on the growth patterns and selection of local varieties of sugi (Cryptomeria japonica).
We analyzed the effect of genotype and planting density on the growth patterns of height and diameter at breast height (DBH) and the efficiency of indirect early selection targeting on the low planting density using stem analysis data obtained from a test site composed of 6 cultivars and 3 planting densities (5,000, 3,000, 1,500 trees/ha) at stand age 28. The growth pattern of height was mainly affected by genotype, but the effect of planting density was small. The growth pattern of DBH was affected by both genotype and planting density. Our results showed the importance of selection of the genotype of seedlings for the control of growth in plantations of Sugi. The results of genetic evaluation obtained at general planting density (3,000 trees/ha) could be applied for the prediction of genetic performance of growth traits at different planting densities. Approximately 15 years would be required to obtain 80％ of genetic gain at stand age 28 with the plantation of 1,500 trees/ha according to the test results obtained at the plantation of general planting density (3,000 trees/ha).

DOI： 10.4005/jjfs.98.45

Language：Japanese  

Application of Terrestrial LiDAR for Forest Tree Breeding: Evaluation of Stem Traits on F1 Progenies of Cryptomeria japonica Plus Trees.
In order to apply terrestrial LiDAR for phenotyping in forest tree breeding, various traits of stem were quantified and evaluated based on a point cloud obtained by ground-based three-dimensional laser measurements on a study site of F₁ individuals of Cryptomeria japonica plus trees. The point cloud of each individual stem was approximated by "stem circles", which are horizontal perfect circles arrayed per 10 cm height. Based on the point cloud and the stem circles, both height and diameter at breast height (DBH) of individuals were automatically estimated with high accuracy. The relative stem taper curve equation of the third order was approximated for each individual based on the stem circles. Stem volume of each individual estimated from the taper curve showed high correlation with one estimated from only tree height and DBH. Stem straightness was quantified in the two standardized methods and these methods were evaluated with stem straightness index investigated on the field. The relationships among these estimated traits were examiend using correlation coefficients. Of the ratio of additive genetic variance, the highest was stem straightness, followed in order by tree height, stem volume, DBH, and the coefficients of the relative stem taper curve. The usefulness of the analytical method for phenotyping in forest tree breeding was discussed.

DOI： 10.4005/jjfs.97.290

Language：Others   Publishing type：Research paper (scientific journal)  

Conifers have larger genomes than most angiosperms, long generation times, and undergone relatively few chromosome duplications during their evolution. Thus, conifers are interesting targets for molecular evolutionary studies. Despite this, there have been few studies regarding their genome structure, and these studies are mostly limited to the Pinaceae. Our target species, Cryptomeria japonica, belongs to the Cupressaceae family, which is phylogenetically separated from the Pinaceae family by a few hundred million years, and is the most important timber tree in Japan, making investigation of its genome structure both interesting and worthwhile. We analyzed the sequences of eight random bacterial artificial chromosome (BAC) clones from C. japonica and compared them with sequences of comparable size from eight other model plants, including Arabidopsis thaliana and Pinus taeda. From this analysis, we identified several features of the C. japonica genome. First, the genome of C. japonica has many divergent repetitive sequences, similar to those of Physcomitrella patens and P. taeda. Additionally, some C. japonica transposable elements (TEs) seem to have been active until recently, and some might be unidentified novel TEs. We also found a putative protein-coding gene with a very long intron (approximately 70 kb). The three Pinaceae species whose genome sequences have been determined share these features, despite the few hundred million years of independent evolution separating the Pinaceae species from C. japonica.

DOI： 10.1007/s11295-015-0859-9

Language：Others   Publishing type：Research paper (scientific journal)  

DOI： 10.1007/s10310-015-0489-y

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1007/s10310-014-0477-7

Language：Others   Publishing type：Research paper (scientific journal)  

The Yezo spruce (Picea jezoensis) has attracted attention as a species commonly used for afforestation in Hokkaido. However, there is no information available for assessing appropriate seed and planting zones for this species. We developed 21 nuclear microsatellite markers for P. jezoensis and used them to examine the genetic structure of nine natural populations (288 trees). Based on the 31 samples, the 21 markers were highly polymorphic, with an average expected heterozygosity greater than 0.84. When we applied ten effective markers to nine populations in Hokkaido, no clear regional differences in genetic diversity were detected. STRUCTURE analysis revealed that, aside from certain geographically isolated or peripheral populations, the populations within planting zones belonged to a single cluster. Thus, based on our marker data, with a few exceptions, there appears to be a single seed zone within the planting zones in Hokkaido.

DOI： 10.1007/s10310-014-0477-7

Language：English   Publishing type：Research paper (scientific journal)  

Language：Others   Publishing type：Research paper (scientific journal)  

Teak (Tectona grandis), which is among the most valuable tropical timber species, is under pressure from rapid deforestation and habitat fragmentation. Limited genetic information is available for Myanmar teak, which comes from the largest natural teak-bearing forest area in the world. To determine the phylogeographic patterns of Myanmar teak, we evaluated three newly developed chloroplast single nucleotide polymorphism (cpSNP) and 10 nuclear simple sequence repeat (nrSSR) markers in 480 individuals representing 20 natural populations. The cpSNP markers detected four haplotypes, each differing by a single mutation. The GST value was lower than the NST value and did not reveal a phylogeographic structure of Myanmar teak. Nuclear microsatellite analysis revealed high genetic diversity with the mean expected heterozygosity (0.652). The same level of genetic differentiation (4%) was observed for both cpSNP and nrSSR markers in different groups. Conservation of the HMB, TDG, KTA, and POL populations should receive highest priority because these contribute most to the total genetic diversity. The genetic boundaries of teak observed from combining the results of cpSNP and nrSSR marker barriers suggested four potential zones in the teak seed transfer guidelines of Myanmar. In light of our findings, we discussed appropriate gene conservation strategies and proposed seed zones to safeguard the current genetic resources of Myanmar teak.

DOI： 10.1177/194008291500800118

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1007/s11295-014-0758-5

Language：Others   Publishing type：Research paper (scientific journal)  

Population structure and genome-wide linkage disequilibrium (LD)were investigated in a Cryptomeria japonica core collection consisting of 510 individuals and providing comprehensive coverage of the natural distribution range of the species, using 4,896 SNP markers. Both Bayesian clustering and principal coordinate analyses clearly subdivided the sample into two distinct groups corresponding to the Japan Sea side and the Pacific Ocean side of Japan. The clustering analysis also detected two subgroups located in Northern Tohoku and Yakushima Island, as was also the case in the natural population. This finding is in accord with the ecogeographical origin of the cultivars; however, the spatial distribution of these clusters was slightly different between the natural population and the core collection, especially in the Chugoku district, an effect which might have resulted from historical human activity. The genetic diversity of the core collection is comparable to, or a little higher than, that of natural populations, indicating that the individuals in the core collection have not suffered from diversity losses caused by a domestication bottleneck. LD analysis with mapped markers indicated that the LD decays rapidly, within approximately 5 cM, in both the core collection and natural populations. The results show that the population described here is a valuable resource for both basic and applied research on C. japonica.

DOI： 10.1007/s11295-014-0758-5

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• Context: Little is known about the potential of gene flow and resulting genetic structures of trees employing sea-drifting seed dispersal in island populations. • Aims: Current genetic structure and the magnitude of historical gene flow were estimated in island populations of Calophyllum inophyllum L., a typical plant employing sea-drifting seed dispersal. • Methods: Samples were collected from the northern extreme of the species' distribution (Taiwan and the Sakishima, Daito, and Ogasawara Islands, Japan) and genotyped using 15 EST-SSR markers. Genetic differentiation (F ST and AMOVA), genetic structure (STRUCTRE analysis), and historical gene flow (assignment testing) were determined. • Results: Frequent gene flow within and rare gene flow among island groups was determined using assignment testing. Clear genetic structures were also detected using the STRUCTURE analysis, which demonstrated differentiation between dominant clusters among geographically constructed island groups. • Conclusions: The potential for gene flow via sea-drifting seed dispersal was high, and this was possible even among small islands. However, the extent and frequency of gene flow were not great enough to prevent genetic differentiation in a range of over a few hundred kilometers. © 2014 INRA and Springer-Verlag France.

DOI： 10.1007/s13595-014-0365-5

Language：English   Publishing type：Research paper (scientific journal)  

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Pine wilt disease caused by the pine wood nematode (PWN), Bursaphelenchus xylophilus, has been epidemic and has had disastrous impacts on pine forests and forest ecosystems in eastern Asia. Many pine species in this area are susceptible to this disease. Pinus thunbergii is particularly susceptible. In Japan, tree breeders have selected surviving trees from severely damaged forests as resistant candidates, and have finally established several resistant varieties of P. thunbergii. However, this breeding procedure requires much time and effort due to the lack of physiological and phenotypical information about resistance. To investigate the resistance mechanisms of selected P. thunbergii, we compared histochemical responses, tissue damage expansion, and PWN distribution in resistant and susceptible clones of P. thunbergii after PWN inoculation. The results suggested that the mechanisms of resistance are as follows: damage expansion in the cortex, cambium, and xylem axial resin canals are retarded in resistant trees soon after inoculation, probably due to the induction of wall protein-based defenses. Suppression of PWN reproduction was particularly caused by inhibition of damage expansion in the cambium. The slow expansion of damage in each tissue provides time for the host to complete the biosynthesis of lignin in the walls of cells that surround the damaged regions. This lignification of cell walls is assumed to effectively inhibit the migration and reproduction of the PWNs. The mechanism of initial damage retardation is presumed to be a key for resistance. © 2013 The Author(s).

DOI： 10.1007/s10310-013-0417-y

Language：English   Publishing type：Research paper (scientific journal)  

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Background: Forest trees have ecological and economic importance, and Japanese cedar has highly valued wood attributes. Thus, studies of molecular aspects of wood formation offer practical information that may be used for screening and forward genetics approaches to improving wood quality.Results: After identifying expressed sequence tags in Japanese cedar tissue undergoing xylogenesis, we designed a custom cDNA microarray to compare expression of highly regulated genes throughout a growing season. This led to identification of candidate genes involved both in wood formation and later cessation of growth and dormancy. Based on homology to orthologous protein groups, the genes were assigned to functional classes. A high proportion of sequences fell into functional classes related to posttranscriptional modification and signal transduction, while transcription factors and genes involved in the metabolism of sugars, cell-wall synthesis and lignification, and cold hardiness were among other classes of genes identified as having a potential role in xylem formation and seasonal wood formation.Conclusions: We obtained 55,051 unique sequences by next-generation sequencing of a cDNA library prepared from cambial meristem and derivative cells. Previous studies on conifers have identified unique sequences expressed in developing xylem, but this is the first comprehensive study utilizing a collection of expressed sequence tags for expression studies related to xylem formation in Japanese cedar, which belongs to a different lineage than the Pinaceae. Our characterization of these sequences should allow comparative studies of genome evolution and functional genetics of wood species. © 2014 Mishima et al.; licensee BioMed Central Ltd.

DOI： 10.1186/1471-2164-15-219

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In order to improve the accessibility of genetic resources, a core collection needs to be constructed and appropriately evaluated. For Cryptomeria japonica, the most important forestry species in Japan, we constructed a core collection derived from 3,203 plus trees. First, genetic redundancy was removed based on SSR genotypes from the plus tree population. One diploid accession from each of the 539 groups that were identified using information on the original growing location was then selected considering the conservation conditions. We defined this population composed of 539 individuals as a core collection for C. japonica, and then evaluated the collection status of this core collection from multiple viewpoints. This core collection proved to contain on average one entry selected per 10 km radius (314 km2) from 10 × 10 km grid squares with an area of more than 40 % covered with C. japonica, and covered more than 85 % retention of all 12 environmental factors affecting plus trees, while maintaining almost the same genetic diversity as the complete set of plus trees. These methods of evaluation from multiple viewpoints, using a geographic information system and genetic markers, were efficient in determining whether the collection of resources was complete, and, if not, where additional efforts for balancing a deficit should be devoted.

DOI： 10.1007/s10310-014-0460-3

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Genome-wide association studies (GWAS) are an alternative to bi-parental QTL mapping in long-lived perennials. In the present study, we examined the potential of GWAS in conifers using 367 unrelated plus trees of Cryptomeria japonica D. Don, which is the most widely planted and commercially important tree species in Japan, and tried to detect significant associations between wood property traits and quantity of male strobili on the one hand, and 1,032 single nucleotide polymorphisms (SNPs) assigned to 1,032 genes on the other. Association analysis was performed with the mixed linear model taking into account kinship relationships and subpopulation structure. In total, 6 SNPs were found to have significant associations with the variations in phenotype. These SNPs were not associated with the positions of known genes and QTLs that have been reported to date, thus they may identify novel QTLs. These 6 SNPs were all found in sequences showing similarities with known genes, although further analysis is required to dissect the ways in which they affect wood property traits and abundance of male strobili. These presumptive QTL loci provide opportunities for improvement of C. japonica, based on a marker approach. The results suggest that GWAS has potential for use in future breeding programs in C. japonica. © 2013 Uchiyama et al.

DOI： 10.1371/journal.pone.0079866

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Long-distance dispersal (LDD) of seeds has a critical impact on species survival in patchy landscapes. However, relative to pollen dispersal, empirical data on how seed LDD affects genetic diversity in fragmented populations have been poorly reported. Thus, we attempted to indirectly evaluate the influence of seed LDD by estimating maternal and paternal inbreeding in the seed rain of fragmented 8 Pinus densiflora populations. In total, the sample size was 458 seeds and 306 adult trees. Inbreeding was estimated by common parentage analysis to evaluate gene flow within populations and by sibship reconstruction analysis to estimate gene flow within and among populations. In the parentage analysis, the observed probability that sampled seeds had the same parents within populations was significantly larger than the expected probability in many populations. This result suggested that gene dispersal was limited to within populations. In the sibship reconstruction, many donors both within and among populations appeared to contribute to sampled seeds. Significant differences in sibling ratios were not detected between paternity and maternity. These results suggested that seed-mediated gene flow and pollen-mediated gene flow from outside population contributed some extent to high genetic diversity of the seed rain (HE > 0.854). We emphasize that pine seeds may have excellent potential for gene exchange within and among populations. © The American Genetic Association. 2013. All rights reserved.

DOI： 10.1093/jhered/est022

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Pinus kesiya Royle ex Gordon (P. kesiya) is one of the economically and ecologically important pine species in South-East Asian. 12 polymorphic tetranucleotide microsatellite markers were isolated from this species using a dual-suppression polymerase chain reaction (PCR) technique and these markers were characterized in 24 individuals collected from a natural population in the Southern Vietnam. The number of alleles per locus ranged from 2 to 28, and expected heterozygosities of the 12 loci ranged from 0. 198 to 0. 954, with an average of 0. 685. These polymorphic markers will serve as useful tools for studies of conservation genetics in P. kesiya and probably in its closely related species. © 2012 Springer Science+Business Media Dordrecht.

DOI： 10.1007/s12686-012-9814-3

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Pinus merkusii is an important tree native to Southeast Asia. Thirteen tetranucleotide microsatellite makers were developed from this species using a dual-suppression polymerase chain reaction technique. Of these markers, nine loci were polymorphic. The mean number of alleles per locus was 3. 18 (range 2-5) and mean expected heterozygosities were 0. 525 (range 0. 384-0. 712). These microsatellite markers will be available to assess the genetic diversity, mating patterns and population structure which will facilitate conservation and management strategies. © 2012 Springer Science+Business Media Dordrecht.

DOI： 10.1007/s12686-012-9821-4

Language：English   Publishing type：Research paper (scientific journal)  

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Key message: GUS analysis inCryptomeria japonicarevealed that theCjMALE1promoter is activated in the male strobilus ofC. japonica. Toward the development of male sterile technology for Cryptomeria japonica, a male strobilus-dominant promoter of C. japonica was isolated. The CjMALE1 gene was isolated from a male strobilus-specific suppression subtractive hybridization (SSH) library, and the promoter was isolated by the TAIL-PCR method. To characterize the CjMALE1 promoter, β-glucuronidase (GUS)-fused genes were constructed and introduced into C. japonica using Agrobacterium tumefaciens. GUS expression from CjMALE1-2. 5 K (2,718 bp fragment)::GUSC. japonica and CjMALE1-1 K (1,029 bp fragment)::GUSC. japonica was detected in the tapetum and microspore mother cells. These promoter fragments were comparably active in the pre-meiotic stage of the male strobilus of C. japonica. Our analysis showed that the 1,029 bp promoter had all the cis-elements necessary for male strobilus-dominant expression of CjMALE1. When CjMALE1-1 K::GUS was introduced into Arabidopsis, GUS expression was detected in the same spatiotemporal pattern as in C. japonica. These results suggest that the CjMALE1 promoter is subject to transcriptional regulatory systems consisting of cis- and trans-elements that have been highly conserved during evolution. © 2012 Springer-Verlag Berlin Heidelberg.

DOI： 10.1007/s00299-012-1365-2

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

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We attempted to evaluate the genetic diversity of long-distance transported pollen flowing over fragmented Pinus densiflora populations during a mating season. A P. densiflora clonal seed orchard, which was located in a rural area where many fragmented populations exist, was selected for pollen capture. Immigrant pollen captured by three clones having different flowering times was regarded as the pollen flowing over fragmented populations during a mating season. The genetic diversity (He) values of the immigrant pollen captured by the three clones were high (He > 0.894). The correlation of paternity (rp) values of the seeds having immigrant parent generated from the three clones were calculated to be negative. From these parameters, the pollen cloud is considered to have maintained high genetic diversity during the mating season. The genetic composition of the pollen cloud showed slight variation. The pollen captured by different trees (i. e., clonal ramets of the three clones) was significantly different based on analysis of molecular variance. Especially, the pollen pools captured by trees planted in the western side of the orchard were significantly different from the gene pool of the surrounding populations. Factors affecting this differentiation could be that the donors of the pollen transported to the orchard vary with time, as well as nonuniform dispersal of the pollen. From these results, the pollen flowing over fragmented P. densiflora populations is considered to have high genetic diversity, compensating to some extent for fragmentation. © 2011 The Japanese Forest Society and Springer.

DOI： 10.1007/s10310-011-0322-1

Language：English   Publishing type：Research paper (scientific journal)  

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The genetic differentiation inherent in Zelkova serrata in Japan and the southern portion of the Korean Peninsula was examined by comparing a chloroplast DNA (cpDNA) sequence over a 16 k baselength in 40 individual samples collected from an area covering the natural distribution range of Z. serrata in Japan and the southern portion of the Korean Peninsula. We detected over 50 single-nucleotide polymorphisms in the protein-coding and intergenic regions, and over 30 insertions/deletions in the intergenic region. From the polymorphisms detected in the cpDNA, 14 haplotypes were identified. These 14 haplotypes had cluster-like structures and genetic differentiation between the clusters was large. Closely related haplotypes existed in adjacent regions. One haplotype existed in both Japan and the Korean Peninsula. By comparison with other Zelkova species, Z. serrata is apparently distinct from European and East Asian Zelkova species and Z. serrata is closest to the Ulmus species in the genus Zelkova. The effects of the analyzed length of the cpDNA sequence on the detection of polymorphisms were analyzed by re-sampling simulation. © 2012 Springer Science+Business Media B.V.

DOI： 10.1007/s10592-012-0358-6

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Premise of the study: Microsatellite markers were developed for the endemic Japanese species Thujopsis dolabrata var. hondai for studies on forest ecology and tree breeding. Methods and Results: We characterized 10 dinucleotide microsatellite loci by screening primers developed using a simple sequence repeat-enriched library. The number of alleles per locus ranged from eight to 44 with an average of 20.3. The observed and expected heterozygosities ranged from 0.326 to 0.854 and from 0.670 to 0.976, respectively. Conclusions: The development of these markers can be used to assess useful genetic information for ecological studies and tree breeding in T. dolabrata var. hondai. © 2012 Botanical Society of America.

DOI： 10.3732/ajb.1200010

Language：English   Publishing type：Research paper (scientific journal)  

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Melia volkensii Gurke is a drought-tolerant tree endemic to the drylands of eastern Africa. In Kenya, the tree has been overexploited for use as timber, and domestication efforts have demonstrated diversity in both genetically controlled tree growth and form. Genetic markers, however, have not yet been developed to support conservation and breeding efforts in the species. In the present study, 15 novel microsatellite markers were developed for M. volkensii Gurke. Genetic variation at these loci was examined in 89 samples from three natural populations in Kenya. All loci were highly polymorphic with mean number of alleles, expected heterozygosity and observed heterozygosity per locus in samples collected from three natural populations in Kenya having values of 9.92, 0.72 and 0.67, respectively. The microsatellite markers developed in the present study will serve as useful tools in the study of population genetics, ecology, conservation and management of M. volkensii. © 2011 Springer Science+Business Media B.V.

DOI： 10.1007/s12686-011-9558-5

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

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Language：Others   Publishing type：Research paper (scientific journal)  

In order to examine whether seedlings of non-native genetic lineages had been planted, we compared chloroplast DNA haplotypes in plantations and neighboring natural populations of Fagus crenata in Nagano Prefecture, Japan. This region was chosen for study because there are abundant natural F. crenata populations with different haplotypes, and because the species is also frequently used for afforestation in the area. We sampled 159 trees from 30 populations of the species across most of its natural range in the region, and 136 trees from 20 plantations of the species, ranging in age from 3 to 17 years. Six single nucleotide polymorphisms and four insertions/deletions in two chloroplast DNA regions (trnL-trnF and trnK) were analyzed in each of the sampled individuals in order to determine their haplotypes. Four haplotypes (B, D, E and F) were detected in the natural populations, and these exhibited a clear geographical structure; in contrast, only two haplotypes (A and B) were found in the plantations. We found that 15 out of the 20 plantations-located in central and southern areas on the Pacific side of Honshu, where the natural populations contain haplotypes D, E, and F, and where the climate is characterized by dry, cold winters-had been established using seedlings with haplotypes A and B derived from the Sea of Japan side of the Japanese Islands, where it snows heavily. The risks associated with planting seedlings of inappropriate lineages are discussed. © 2011 The Japanese Forest Society and Springer.

DOI： 10.1007/s10310-011-0262-9

Language：Others   Publishing type：Research paper (scientific journal)  

Background: Pine wilt disease is caused by the pine wood nematode, Bursaphelenchus xylophilus, which threatens pine forests and forest ecosystems worldwide and causes serious economic losses. In the 40 years since the pathogen was identified, the physiological changes occurring as the disease progresses have been characterized using anatomical and biochemical methods, and resistant trees have been selected via breeding programs. However, no studies have assessed the molecular genetics, e.g. transcriptional changes, associated with infection-induced physiological changes in resistant or susceptible trees.Results: We constructed seven subtractive suppression hybridization (SSH) cDNA libraries using time-course sampling of trees inoculated with pine wood nematode at 1, 3, or 7 days post-inoculation (dpi) in susceptible trees and at 1, 3, 7, or 14 dpi in resistant trees. A total of 3,299 sequences was obtained from these cDNA libraries, including from 138 to 315 non-redundant sequences in susceptible SSH libraries and from 351 to 435 in resistant SSH libraries. Using Gene Ontology hierarchy, those non-redundant sequences were classified into 15 subcategories of the biological process Gene Ontology category and 17 subcategories of the molecular function category. The transcriptional components revealed by the Gene Ontology classification clearly differed between resistant and susceptible libraries. Some transcripts were discriminative: expression of antimicrobial peptide and putative pathogenesis-related genes (e.g., PR-1b, 2, 3, 4, 5, 6) was much higher in susceptible trees than in resistant trees at every time point, whereas expression of PR-9, PR-10, and cell wall-related genes (e.g., for hydroxyproline-rich glycoprotein precursor and extensin) was higher in resistant trees than in susceptible trees at 7 and 14 dpi.Conclusions: Following inoculation with pine wood nematode, there were marked differences between resistant and susceptible trees in transcript diversity and the timing and level of transcripts expressed in common; in particular, expression of stress response and defense genes differed. This study provided new insight into the differences in the physiological changes between resistant and susceptible trees that have been observed in anatomical and biochemical studies. © 2012 Hirao et al; licensee BioMed Central Ltd.

DOI： 10.1186/1471-2229-12-13

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

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Microsatellite markers were isolated from Robinia pseudoacacia L. using an enrichment method. Eleven of the 23 primer pairs designed successfully amplified unambiguous and polymorphic single loci among 39 individual R. pseudoacacia L. from northeastern Japan. The observed and expected heterozygosities of the 11 microsatellite markers ranged from 0.333 to 0.821 and from 0.489 to 0.867, respectively. The polymorphisms observed at the 11 microsatellite loci are useful genetic data for forest ecological studies involving R. pseudoacacia L. © 2009 Blackwell Publishing Ltd.

DOI： 10.1111/j.1755-0998.2008.02306.x

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

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We developed 17 chloroplast microsatellite markers, which consisted of seven mononucleotide microsatellites with a minimum repeat number of 10 and 10 dinucleotide microsatellites with a minimum repeat number of six, from the complete chloroplast genomic sequence of Cryptomeria japonica. A survey of 45 C. japonica individuals showed the number of alleles ranging from two to 11 alleles and a diversity index ranging from 0.085 to 0.895. Consequently, the 45 C. japonica individuals were divided into 39 haplotypes. These markers will be useful genetic markers in the gene flow analysis and population genetics of C. japonica. © 2008 The Authors.

DOI： 10.1111/j.1755-0998.2008.02216.x

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To investigate how accurately biparentage assignments for coniferous seeds can be improved by using different kinds of seed tissues, we assigned biparentage to dispersed seeds in a natural stand of Pinus densiflora Siebold & Zucc. (288 mature trees) using two procedures: with or without megagametophyte haplotype data ("MH" and "ordinary" procedures, respectively). Using archived clones, we confirmed the conifer-specific modes of inheritance of three kinds of seed tissues from certain maternal trees using six microsatellite markers. In the natural stand, under the MH procedure, 39.2% of male parents and 77.0% of female parents for a total of 204 seeds analyzed were assigned to at least one mature tree within the study area. At that time, the proportion of exactly matching seeds out of seeds with one matching parent under the MH procedure was significantly larger than that of the seeds under the ordinary procedure. The biparentage assignments under the ordinary procedure corresponded to only 53.7% of the accurate separate assignments under the MH procedure. It is suggested that analyzing different seed tissues is effective for exact and accurate biparentage assignments in investigations of biparentally mediated gene flow in coniferous populations, particularly at the seed-dispersal stage. © 2007 NRC.

DOI： 10.1139/X07-046

Language：English   Publishing type：Research paper (scientific journal)  

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Microsatellites were isolated and characterized for the Japanese oak species, Quercus mongolica var. crispula, distributed in temperate deciduous forests of Japan. Eleven of the 48 primer pairs designed successfully amplified unambiguous and polymorphic single loci among 67 Q. mongolica var. crispula individuals within a plot in southwestern Japan. The observed and expected heterozygosities of the 11 microsatellite markers ranged from 0.522 to 0.896 and from 0.536 to 0.882, respectively. These polymorphic microsatellite markers are useful for estimating pollen-mediated gene flow in Q. mongolica var. crispula. © 2006 The Authors.

DOI： 10.1111/j.1471-8286.2006.01313.x

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Microsatellites were isolated and characterized for Japanese larch, Larix kaempferi, a conifer species distributed in Japan. A larch genomic DNA library enriched for (AG)n repeats was screened using the colony polymerase chain reaction method and 145 unique microsatellite containing sequences were obtained. Seventy-two primer pairs were designed and 30 produced single-locus products, and 19 of them were polymorphic. The expected heterozygosity ranged from 0.566 to 0.951. These 19 polymorphic microsatellite loci should be valuable markers for genetic studies on Japanese larch. © 2006 The Authors.

DOI： 10.1111/j.1471-8286.2006.01291.x

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Language：Others   Publishing type：Research paper (scientific journal)  

We developed seven microsatellite loci from Pinus densiflora using a dual polymerase chain reaction (PCR) technique. Of 186 clones from a library based on suppression PCR, 127 contained microsatellite sequences. Of these, 43 candidates were determined sequences of both flanking regions, and 16 regions from this group were chosen as development markers. Seven of these primer pairs successfully amplified polymorphic single loci among 83 resistant trees against pine wood nematode. The observed heterozygosity of the seven microsatellite markers ranged from 0.247 to 0.843. Mendelian inheritance was confirmed using megagametophytes. © 2006 Blackwell Publishing Ltd.

DOI： 10.1111/j.1471-8286.2005.01145.x

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

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We developed 14 microsatellite loci from an enriched genomic DNA library of a broad-leaved deciduous tree, Zelkova serrata. Of 198 clones from the library, 112 contained microsatellite repeat regions. The M13-tailed primer method was used for economy. Sequence-specific primer pairs were designed for 58 of 76 candidate clones. Fourteen of these primer pairs successfully amplified polymorphic single loci among 34 individuals collected from the Kanto breeding region in Japan. The expected heterozygosity for the 14 microsatellite markers ranged from 0.378 to 0.876, suggesting that these will prove valuable for breeding and ecological studies on Z. serrata. © 2005 Blackwell Publishing Ltd.

DOI： 10.1111/j.1471-8286.2005.00933.x

Language：English   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)  

Language：Japanese   Publishing type：Research paper (scientific journal)  

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Language：English   Publishing type：Research paper (scientific journal)  

Language：Japanese   Book type：General book, introductory book for general audience

Responsible for pages：第３部解析手法編第２章オルガネラDNA解析手法，193-224   Language：Japanese   Book type：Scholarly book

Repository Public URL： http://hdl.handle.net/2324/1001482699

Event date： 2019.3

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2018.8 - 2018.6

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Event date： 2016.5

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：九州大学   Country：Japan  

Event date： 2016.5

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：岩手大学   Country：Japan  

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東京大学   Country：Japan  

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：岩手大学   Country：Japan  

Language：Japanese   Presentation type：Oral presentation (general)  

Country：Japan  

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東京大学   Country：Japan  

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東京大学   Country：Japan  

Language：Japanese   Presentation type：Oral presentation (general)  

Venue：東京大学   Country：Japan  

Language：Japanese   Presentation type：Oral presentation (invited, special)  

Venue：西日本短期大学   Country：Japan  

Language：Japanese  

Venue：北海道大学   Country：Japan  

Language：Japanese  

Venue：北海道大学   Country：Japan  

Language：Japanese  

Country：Japan  

Language：Japanese  

Venue：北海道大学   Country：Japan  

Language：English  

Changes in annual transcriptome dynamics of Japanese cedar planted under different climate conditions
[in Japanese]

DOI： 10.11519/jfsc.133.0_424

Language：Japanese  

スギさし木苗の根系成長におけるクローン間変異
[in Japanese]

DOI： 10.11519/jfsc.127.0_285

Language：Japanese  

スギ精英樹におけるゲノムワイドアソシエーション解析とゲノミック予測

DOI： 10.11519/jfsc.127.0_282

Language：Japanese  

DNAマーカーに基づいた次世代優良少花粉スギの効率的スクリーニング
[in Japanese]

DOI： 10.11519/jfsc.126.0_306

Language：Japanese  

環境因子を考慮したスギさし木クローンの成長解析
[in Japanese]

DOI： 10.11519/jfsc.126.0_337

Language：Japanese  

共通系統を用いたスギ初期成長の産地間差の解析
[in Japanese]

DOI： 10.11519/jfsc.126.0_322

Language：Japanese  

スギ精英樹クローンの苗畑における初期成長の評価
[in Japanese]

DOI： 10.11519/jfsc.126.0_321

Language：Japanese  

スギの材質・成長形質に関するQTL解析
[in Japanese]

DOI： 10.11519/jfsc.126.0_307

Language：Japanese  

スギのさし木発根性の季節変動とその要因
[in Japanese]

DOI： 10.11519/jfsc.126.0_335

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Language：Japanese  

優良な形質を併せ持つ雄性不稔スギの作出―雄性不稔スギリソース整備に向けて―
[in Japanese]

DOI： 10.11519/jfsc.125.0_719

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Language：English  

Language：Others  

We measured linkage disequilibrium in mostly noncoding regions of Cryptomeria japonica, a conifer belonging to Cupressaceae. Linkage disequilibrium was extensive and did not decay even at a distance of 100 kb. The average estimate of the population recombination rate per base pair was 1.55×10 -5 and was <1/70 of that in the coding regions. We discuss the impact of low recombination rates in a large part of the genome on association studies. © 2012 by the Genetics Society of America.

DOI： 10.1534/genetics.111.136697

Language：English  

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Language：Japanese  

分子系統地理学的解析を用いた宮城県内におけるブナ苗木の流通許容範囲の推定

DOI： 10.11519/jfsc.117.0.31.0

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Language：Others  

To achieve a highly reliable clone management of teak (Tectona grandis) plus trees, useful DNA molecular markers were surveyed using RAPD analysis and their ability to discriminate among plus tree clones was examined. A primary screening was performed for 120 arbitrary decamer primers, and 24 primers that generated 26 clear and unambiguous fragments were selected. In the secondary screening, the reproducibility of each fragment was investigated by six repetitions of polymerase chain reaction, and 13 fragments found to be most reproducible were finally selected. Evaluation of the discriminatory powers of these fragments suggested that the selected RAPD markers would be useful in the clone management of teak plus trees.

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Language：Japanese   Publishing type：Article, review, commentary, editorial, etc. (scientific journal)  

Type：Competition, symposium, etc. 

Type：Peer review 

Number of peer-reviewed articles in foreign language journals：15

Type：Competition, symposium, etc. 

Type：Competition, symposium, etc. 

Type：Peer review 

Number of peer-reviewed articles in foreign language journals：17

Type：Competition, symposium, etc. 

Type：Academic society, research group, etc. 

Type：Competition, symposium, etc. 

Type：Peer review 

Number of peer-reviewed articles in foreign language journals：17

Number of peer-reviewed articles in Japanese journals：2

Type：Peer review 

Number of peer-reviewed articles in foreign language journals：8

Number of peer-reviewed articles in Japanese journals：1

Type：Competition, symposium, etc. 

Type：Competition, symposium, etc. 

Number of participants：100

Type：Competition, symposium, etc. 

Type：Competition, symposium, etc. 

Number of participants：50

Type：Competition, symposium, etc. 

Type：Competition, symposium, etc. 

Type：Competition, symposium, etc. 

Number of participants：100

Type：Competition, symposium, etc. 

Number of participants：60

Type：Competition, symposium, etc. 

Number of participants：150

Type：Competition, symposium, etc. 

Type：Competition, symposium, etc. 

Number of participants：150

Type：Academic society, research group, etc. 

Audience：Infants,　Schoolchildren,　Junior students,　High school students

Type：Seminar, workshop