Mitochondrial (mt) DNA variation in Japanese beech, Fagus crenata (Fagaceae), was studied in 17 populations distributed throughout the species' range. Total genomic DNA of samples from single trees representing each of 12 populations were digested with 18 restriction enzymes and hybridized with three probes containing coxI, coxIII, and atpA gene sequences. Thirty-four of the 54 enzyme/probe combinations showed polymorphisms and all the individuals were subsequently analyzed with six combinations of three probes and two enzymes. Restriction fragment length polymorphisms were evident around all three genes, allowing the identification of eight distinct haplotypes. Haplotype diversity within the populations was found to be very low (HS = 0.031), but population differentiation to be much higher (GST = 0.963). The mtDNA variation was strikingly different from allozyme variation (HS = 0.209; GST = 0.039). Gene flow for maternally inherited mtDNA should be restricted to seed dispersal while nuclear gene flow occurs by both seed and pollen dispersal. Therefore, the difference in the variation between mtDNA and allozymes may be largely a result of the much higher rate of gene flow associated with pollen dispersal than with seed dispersal. The mtDNA variation displayed strong geographic structure, which may reflect the species' distribution in the last glacial maximum and subsequent colonization, and probably also reflects intraspecific phylogeography of the species.

Tohoku Regional Breeding Of®ce, Forest Tree Breeding Centre, Takizawa, Iwate 020-0173, Japan and àHokkaido Regional Breeding Of®ce, Forest Tree Breeding Centre, Ebetsu, Hokkaido 069-0836, JapanTo examine the eects of forest cutting on within-population genetic structure, the genetic structure and variability of two Japanese beech (Fagus crenata Blume) stands with contrasting histories in relation to cutting were investigated. Six hundred and sixty beech trees, covering two hectares in total, were mapped and genetically analysed using nine isozyme loci encoding eight enzyme systems. The proportion of polymorphic loci, the average number of alleles per locus, the eective number of alleles per locus, the expected heterozygosity and the observed heterozygosity were 78, 3.3, 1.31, 0.200 and 0.189, respectively, in a secondary stand (designated AK) cut during the 1920s. Corresponding ®gures were 78, 3.3, 1.33, 0.203 and 0.193, respectively, in a primary stand designated KU. The inbreeding coecient and the grand mean of the number of alleles in common (NAC) were 0.055 and 1.684 in AK, and 0.042 and 1.649 in KU, respectively. The genetic variability was slightly but signi®cantly lower in AK. The genetic structure of the two stands was strikingly dierent. The proportions of positively signi®cant Moran's I and SND values found in the shortest distance class were 0.86 and 0.38 for AK, and 0.14 and 0.29 for KU, respectively. Furthermore, signi®cant linkage disequilibrium was observed in AK, but none at all in KU. To examine which, if any, dierences in the genetic structure would be likely to in¯uence succeeding generations, we simulated a self-thinning process. The simulation suggested that reduced genetic variability and linkage disequilibrium would have signi®cant in¯uence in the AK stand for several generations.Keywords: Fagus crenata, founder eect, isozyme variability, linkage disequilibrium, self-thinning, spatial autocorrelation. IntroductionForest tree species are genetically more diverse than most other life forms. Generally, the autosomal genetic variation of such species is largely found in the withinpopulation component, and factors such as range of geographical distribution, mating system and seed dispersal all in¯uence the genetic variability retained in the species (Hamrick et al., 1992). As a tool to study how genetic variation is distributed within populations, spatial autocorrelation techniques (Sokal & Oden, 1978a,b) have played an important role in many studies of genetic structure in forest tree species. These techniques are eective for identifying the major genetic processes involved in the generation of genetic structures in speci®c populations (Sokal & Jacquez, 1991; Sokal et al., 1997). Simulation studies and forest tree population studies have revealed that mating system and seed dispersal have considerable in¯uence on within-population genetic structure (Sokal & Wartenberg, 1983). Coniferous species (in which pollination and seed dispersal occurs mainly by wind, and outcrossing rates are high) h...

Genetic diversity at 11 loci encoding nine enzymes was studied in 23 populations of Japanese beech Fagus crenata Blume distributed throughout the range of the species. Levels of genetic diversity were high for both within species (expected mean heterozygosity: 0.194) and within populations (expected mean heterozygosity: 0.187), whereas the level of genetic diversity among populations was low (GsT = 0.038), as observed in various long-lived, woody plants. Despite the low differentiation among populations, geographical patterning of the variation was observed. Populations in southwestern Japan tended to have greater within-population variation and to be more highly differentiated when compared with those in northeastern Japan. In addition, allele frequencies observed at eight loci were significantly related to latitudinal and/or longitudinal gradients and showed clinal variation across the range of the species. Principal components analysis revealed that the populations tended to cluster according to their geographical locations. The nonrandom patterns of variation were probably shaped by relatively recent historical events such as late-Quaternary migration and founding events.

BackgroundJapanese 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.ResultsThis 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.ConclusionsMore 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.Electronic supplementary materialThe online version of this article (10.1186/s12864-018-4581-5) contains supplementary material, which is available to authorized users.

Genetic diversity at 11 loci encoding nine enzymes was studied in 23 populations of Japanese beech Fagus crenata Blume distributed throughout the range of the species. Levels of genetic diversity were high for both within species (expected mean heterozygosity: 0.194) and within populations (expected mean heterozygosity: 0.187), whereas the level of genetic diversity among populations was low ( GsT = 0.038), as observed in various long-lived, woody plants.Despite the low differentiation among populations, geographical patterning of the variation was observed. Populations in south-western Japan tended to have greater within-population variation and to be more highly differentiated when compared with those in north-eastern Japan. In addition, allele frequencies observed at eight loci were significantly related to latitudinal and/or longitudinal gradients and showed clinal variation across the range of the species. Principal components analysis revealed that the populations tended to cluster according to their geographical locations. The nonrandom patterns of variation were probably shaped by relatively recent historical events such as late-Quaternary migration and founding events.