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European hazelnut (Corylus avellana L.) is an important nut crop due to its nutritional benefits, culinary uses, and economic value. Türkiye is the leading producer of hazelnut, followed by Italy and the United States. Quantitative trait locus studies offer promising opportunities for breeders and geneticists to identify genomic regions controlling desirable traits in hazelnut. A genome-wide association analysis was conducted with 5,567 single nucleotide polymorphisms on a Turkish core set of 86 hazelnut accessions, revealing 189 quantitative trait nucleotides (QTNs) associated with 22 of 31 traits (p < 2.9E-07). These QTNs were associated with plant and leaf, phenological, reproductive, nut, and kernel traits. Based on the close physical distance of QTNs associated with the same trait, we identified 23 quantitative trait loci. Furthermore, we identified 23 loci of multiple QTs comprising chromosome locations associated with more than one trait at the same position or in close proximity. A total of 159 candidate genes were identified for 189 QTNs, with 122 of them containing significant conserved protein domains. Some candidate matches to known proteins/domains were highly significant, suggesting that they have similar functions as their matches. This comprehensive study provides valuable insights for the development of breeding strategies and the improvement of hazelnut and enhances the understanding of the genetic architecture of complex traits by proposing candidate genes and potential functions.

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In the face of climate change, bringing more useful alleles and genes from wild relatives of wheat is crucial to develop climate-resilient varieties. We used two populations of backcrossed recombinant inbred lines (BIL1 and BIL2), developed by crossing and backcrossing two intra-specifically diverse Aegilops tauschii accessions from lineage 1 and lineage 2, respectively, with the common wheat cultivar 'Norin 61'. This study aimed to identify quantitative trait loci (QTLs) associated with heat stress (HS) tolerance. The two BILs were evaluated under heat stress environments in Sudan for phenology, plant height (PH), grain yield (GY), biomass (BIO), harvest index (HI), and thousand-kernel weight (TKW). Grain yield was significantly correlated with BIO and TKW under HS; therefore, the stress tolerance index (STI) was calculated for these traits as well as for GY. A total of 16 heat-tolerant lines were identified based on GY and STI-GY. The QTL analysis performed using inclusive composite interval mapping identified a total of 40 QTLs in BIL1 and 153 QTLs in BIL2 across all environments. We detected 39 QTLs associated with GY-STI, BIO-STI, and TKW-STI in both populations (14 in BIL1 and 25 in BIL2). The QTLs associated with STI were detected on chromosomes 1A, 3A, 5A, 2B, 4B, and all the D-subgenomes. We found that QTLs were detected only under HS for GY on chromosome 5A, TKW on 3B and 5B, PH on 3B and 4B, and grain filling duration on 2B. The higher number of QTLs identified in BIL2 for heat stress tolerance suggests the importance of assessing the effects of intraspecific variation of Ae. tauschii in wheat breeding as it could modulate the heat stress responses/adaptation. Our study provides useful genetic resources for uncovering heat-tolerant QTLs for wheat improvement for heat stress environments.

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BACKGROUND: Moso bamboo (Phyllostachys edulis) is a typical East Asian bamboo that does not flower for > 60 years and propagates without seed reproduction. Thus, Moso bamboo can be propagated vegetatively, possibly resulting in highly heterozygous genetic inheritance. Recently, a draft genome of Moso bamboo was reported, followed by whole genome single nucleotide polymorphisms (SNP) analysis, which showed that the genome of Moso bamboo in China has regional characteristics. Moso bamboo in Japan is thought to have been introduced from China over the sea in 1736. However, it is unclear where and how Moso bamboo was introduced in Japan from China. Here, based on detailed analysis of heterozygosity in genome diversity, we estimate the spread of genome diversity and its pedigree of Moso bamboo. RESULTS: We sequenced the whole genome of Moso bamboo in Japan and compared them with data reported previously from 15 regions of China. Only 4.1 million loci (0.37% of the analyzed genomic region) were identified as polymorphic loci. We next narrowed down the number of polymorphic loci using several filters and extracted more reliable SNPs. Among the 414,952 high-quality SNPs, 319,431 (77%) loci were identified as heterozygous common to all tested samples. The result suggested that all tested samples were clones via vegetative reproduction. Somatic mutations may accumulate in a heterozygous manner within a single clone. We examined common heterozygous loci between samples from Japan and elsewhere, from which we inferred that an individual closely related to the sample from Fujian, China, was introduced to Japan across the sea without seed reproduction. In addition, we collected 16 samples from four nearby bamboo forests in Japan and performed SNP and insertion/deletion analyses using a genotyping by sequencing (GBS) method. The results suggested that a small number of somatic mutations would spread within and between bamboo groves. CONCLUSIONS: High heterozygosity in the genome-wide diversity of Moso bamboo implies the vegetative propagation of Moso bamboo from China to Japan, the pedigree of Moso bamboo in Japan, and becomes a useful marker to approach the spread of genome diversity in clonal plants.

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The genetic dissection of agronomically important traits in closely related Japanese rice cultivars is still in its infancy mainly because of the narrow genetic diversity within japonica rice cultivars. In an attempt to unveil potential polymorphism between closely related Japanese rice cultivars, we used a next-generation-sequencing-based genotyping method: genotyping by random amplicon sequencing-direct (GRAS-Di) to develop genetic linkage maps. In this study, four recombinant inbred line (RIL) populations and their parents were used. A final RIL number of 190 for RIL71, 96 for RIL98, 95 for RIL16, and 94 for RIL91 derived from crosses between a common leading Japanese rice cultivar Koshihikari and Yamadanishiki, Taichung 65, Fujisaka 5, and Futaba, respectively, and the parent plants were subjected to GRAS-Di library construction and sequencing. Approximately 438.7 Mbp, 440 Mbp, 403.1 Mbp, and 392 Mbp called bases covering 97.5%, 97.3%, 98.3%, and 96.1%, respectively, of the estimated rice genome sequence at average depth of 1× were generated. Analysis of genotypic data identified 1050, 1285, 1708, and 1704 markers for each of the above RIL populations, respectively. Markers generated by GRAS-Di were organized into linkage maps and compared with those generated by GoldenGate SNP assay of the same RIL populations; the average genetic distance between markers showed a clear decrease in the four RIL populations when we integrated markers of both linkage maps. Genetic studies using these markers successfully localized five QTLs associated with heading date on chromosomes 3, 6, and 7 and which previously were identified as Hd1, Hd2, Hd6, Hd16, and Hd17. Therefore, GRAS-Di technology provided a low cost and efficient genotyping to overcome the narrow genetic diversity in closely related Japanese rice cultivars and enabled us to generate a high density linkage map in this germplasm.

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Fusarium wilt is a significant disease in radish, but the genetic mechanisms controlling yellows resistance (YR) are not well understood. This study aimed to identify YR-QTLs and to fine-map one of them using F2:3 populations developed from resistant and susceptible radish parents. In this study, two high-density genetic maps each containing shared co-dominant markers and either female or male dominant markers that spanned 988.6 and 1127.5 cM with average marker densities of 1.40 and 1.53 cM, respectively, were generated using Genotyping by Random Amplicon Sequencing-Direct (GRAS-Di) technology. We identified two YR-QTLs on chromosome R2 and R7, and designated the latter as ForRs1 as the major QTL. Fine mapping narrowed down the ForRs1 locus to a 195 kb region. Among the 16 predicted genes in the delimited region, 4 genes including two receptor-like protein and -kinase genes (RLP/RLK) were identified as prime candidates for ForRs1 based on the nucleotide sequence comparisons between the parents and their predicted functions. This study is the first to use a GRAS-Di for genetic map construction of cruciferous crops and fine map the YR-QTL on the R7 chromosome of radish. These findings will provide groundbreaking insights into radish YR breeding and understanding the genetics of YR mechanism.

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To improve the dairy sector in Cambodia in the future, we aimed to reveal the genetic variation and the milk production in Cambodian crossbred dairy cattle. We calculated the percent (%) milk fat content and the average milk yield per cow (L/day) for two farms (Farm R and M) based on the farmers' records and interviews. The crossbred cows originated from Cambodian local farmers and Thailand breeders in Farm R, whereas the crossbred cows originated in Thailand breeders in Farm M. Then, we performed genetic characterization for 75 individuals from the two farms and an individual Japanese pure Holstein-Friesian cow based on 133,705 single nucleotide polymorphisms (SNPs) obtained by the GRAS-Di method. The milk fat contents in the bulk milk in the dry season and the average milk yield per cow on Farm R were 3.77 ± 0.98% and 7.81 ± 2.66 L/day, respectively, and were higher than those on Farm M (3.35 ± 0.54% and 6.5-7.5 L/day). Cattle originating in Cambodia in Farm R possessed a unique genetic character different from cattle from Thailand in Farm M. The present study suggests that the differences in milk fat content between the two farms might be explained by the genetic differences in crossbred cows.

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BACKGROUND: Common buckwheat is considered a quantitative short-day plant and is classified into the autumn (highly photoperiod sensitive), summer (weakly photoperiod sensitive), and intermediate ecotype. Understanding ecotype differentiation is essential for adaptive expansion and maximizing yield. The genetic analysis for ecotype has focused on photoperiod-dependent flowering time, whereas post-flowering traits such as seed set and maturity time might also regulate ecotype differentiation. RESULTS: A field experiment revealed that ecotype differentiation is mainly defined by the timing of seed set and maturation, whereas flowering time is less relevant. Thus, we focused on maturity time as a trait that defines the ecotype. To detect QTLs for maturity time, we developed two F2 populations derived from early × late-maturing accessions and intermediate × late-maturing accessions. Using genotyping by random amplicon sequencing-direct analysis, we generated a high-density linkage map. QTL analysis detected two major QTLs for maturity time, one in each F2 population. We also detected QTLs for flowering time at loci different from maturity time QTLs, which suggests that different genetic mechanisms regulate flowering and maturity. Association analysis showed that both QTLs for maturity time were significantly associated with variations in the trait across years. CONCLUSIONS: Maturity time appeared to be more suitable for explaining ecotype differentiation than flowering time, and different genetic mechanisms would regulate the timing of flowering and maturation. The QTLs and QTL-linked markers for maturity time detected here may be useful to extend the cultivation area and to fine-tune the growth period to maximize yield in buckwheat.

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The Tsushima leopard cat (Prionailurus bengalensis euptilurus) (TLC) is a regional population of the Amur leopard cat (P. bengalensis euptilurus) that lives only on the Tsushima Island in Japan and is threatened with extinction. Because the TLC population is small, genetic management is important. In this study, we obtained the draft genome of the TLC and identified single-nucleotide polymorphism (SNP) markers using a genotyping by random amplicon sequencing-direct (GRAS-Di) analysis. We genotyped 31 captive individuals and 50 wild individuals, of which 48 were from a previous study. The identified SNPs were used to clarify the genetic diversity and genetic structure of the wild and captive populations of the TLC. The size of the genome was estimated to be about 2.42 Gb. The number of SNP markers developed was 139, and although PID and probability of exclusion obtained using these SNP markers were not as high as those reported in the studies of other wild species, these SNP markers could be used to identify individuals and parentage. Moreover, the genetic diversity indices of the captive population were similar to those of the wild population. These SNP markers will be useful for understanding the ecology of the TLC and planning conservation strategies.

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Smut disease of sugarcane causes considerable yield losses and the use of resistant varieties is the best control practice. Our group identified a Japanese wild sugarcane with highly smut disease resistance named 'Iriomote8'. In this study, we conducted QTL analysis for smut disease resistance using a mapping population derived from a resistant variety 'Yaenoushie', in which resistance is inherited from 'Iriomote8'. We identified 4813 non-redundant markers using GRAS-Di technology and developed a linkage map of mapping parents. We evaluated smut disease resistance of the mapping population by the inoculation test. Consequently, a large number of clones did not show the disease symptoms and the distribution of smut disease incidence tended to be "L shaped". Composite interval mapping detected an identical QTL for indices of smut disease incidence with a markedly high LOD score (26.6~45.6) at the end of linkage group 8 of 'Yaenoushie'. This QTL explained approximately 50% of the cases of smut disease incidence. In the mapping population, there were no correlations between the indices of smut disease incidence and other agronomic traits. In conclusion, this QTL could be used for marker-assisted selection to significantly improve smut disease resistance without negative effects on other agronomic traits.

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How early stages of speciation in free-spawning marine invertebrates proceed is poorly understood. The Western Pacific abalones, Haliotis discus, H. madaka, and H. gigantea, occur in sympatry with shared breeding season and are capable of producing viable F1 hybrids in spite of being ecologically differentiated. Population genomic analyses revealed that although the three species are genetically distinct, there is evidence for historical and ongoing gene flow among these species. Evidence from demographic modeling suggests that reproductive isolation among the three species started to build in allopatry and has proceeded with gene flow, possibly driven by ecological selection. We identified 27 differentiation islands between the closely related H. discus and H. madaka characterized by high FST and dA, but not high dXY values, as well as high genetic diversity in one H. madaka population. These genomic signatures suggest differentiation driven by recent ecological divergent selection in presence of gene flow outside of the genomic islands of differentiation. The differentiation islands showed low polymorphism in H. gigantea, and both high FST, dXY, and dA values between H. discus and H. gigantea, as well as between H. madaka and H. gigantea. Collectively, the Western Pacific abalones appear to occupy the early stages speciation continuum, and the differentiation islands associated with ecological divergence among the abalones do not appear to have acted as barrier loci to gene flow in the younger divergences but appear to do so in older divergences.

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The novel non-targeted PCR-based genotyping system, namely Genotyping by Random Amplicon Sequencing, Direct (GRAS-Di), is characterized by the simplicity in library construction and robustness against DNA degradation and is expected to facilitate advancements in genetics, in both basic and applied sciences. In this study, we tested the utility of GRAS-Di for genetic analysis in a cultured population of the tiger pufferfish Takifugu rubripes. The genetic analyses included family structure analysis, genetic map construction, and quantitative trait locus (QTL) analysis for the male precocious phenotype using a population consisting of four full-sib families derived from a genetically precocious line. An average of 4.7 million raw reads were obtained from 198 fish. Trimmed reads were mapped onto a Fugu reference genome for genotyping, and 21,938 putative single-nucleotide polymorphisms (SNPs) were obtained. These 22 K SNPs accurately resolved the sibship and parent-offspring pairs. A fine-scale linkage map (total size: 1,949 cM; average interval: 1.75 cM) was constructed from 1,423 effective SNPs, for which the allele inheritance patterns were known. QTL analysis detected a significant locus for testes weight on Chr_14 and three suggestive loci on Chr_1, Chr_8, and Chr_19. The significant QTL was shared by body length and body weight. The effect of each QTL was small (phenotypic variation explained, PVE: 3.1-5.9%), suggesting that the precociousness seen in the cultured pufferfish is polygenic. Taken together, these results indicate that GRAS-Di is a practical genotyping tool for aquaculture species and applicable for molecular breeding programs, such as marker-assisted selection and genomic selection.

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The Tsushima leopard cat (Prionailurus bengalensis euptilurus) lives on Tsushima Island in Japan and is a regional population of the Amur leopard cat; it is threatened with extinction. Its genetic management is important because of the small population. We used genotyping by random amplicon sequencing-direct (GRAS-Di) to develop a draft genome and explore single-nucleotide polymorphism (SNP) markers. The SNPs were analyzed using three genotyping methods (mapping de novo, to the Tsushima leopard cat draft genome, and to the domestic cat genome). We examined the genetic diversity and genetic structure of the Tsushima leopard cat. The genome size was approximately 2.435 Gb. The number of SNPs identified was 133-158. The power of these markers was sufficient for individual and parentage identifications. These SNPs can provide useful information about the life of the Tsushima leopard cat and the pairings and for the introduction of founders to conserve genetic diversity with ex situ conservation. We identified that there are no subpopulations of the Tsushima leopard cat. The identifying units will allow for a concentration of efforts for conservation. SNPs can be applied to the analysis of the leopard cat in other regions, making them useful for comparisons among populations and conservation in other small populations.