Fekih Rym, Ishimaru Yohei, Okada Satoshi, Maeda Michihiro, Miyagi Ryutaro, Obana Takahiro, Suzuki Kazuyo, Inamori Minoru, Enoki Hiroyuki, Yamasaki Masanori
Food Resources Education and Research Center, Graduate School of Agricultural Science, Kobe University, Kasai 675-2103, Japan.
Bioscience and Biotechnology Center, Nagoya University, Nagoya 464-8601, Japan.
Plants (Basel). 2023 Feb 17;12(4):929. doi: 10.3390/plants12040929.
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 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 , , , , and . 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.
由于日本水稻品种内遗传多样性狭窄，对亲缘关系密切的日本水稻品种中重要农艺性状进行基因剖析的工作仍处于起步阶段。为了揭示亲缘关系密切的日本水稻品种之间潜在的多态性，我们使用了一种基于新一代测序的基因分型方法：随机扩增子测序直接基因分型（GRAS-Di）来构建遗传连锁图谱。在本研究中，使用了四个重组自交系（RIL）群体及其亲本。RIL71的最终RIL数量为190个，RIL98为96个，RIL16为95个，RIL91为94个，它们分别来自日本常见的主导水稻品种越光与山田锦、台中65、藤坂5和双叶的杂交组合，对这些亲本植株进行GRAS-Di文库构建和测序。分别产生了约438.7 Mbp、440 Mbp、403.1 Mbp和392 Mbp的被检测碱基，平均深度为1×，分别覆盖估计水稻基因组序列的97.5%、97.3%、98.3%和96.1%。对基因型数据的分析分别为上述每个RIL群体鉴定出1050、1285、1708和1704个标记。由GRAS-Di产生的标记被整合到连锁图谱中，并与相同RIL群体的GoldenGate SNP分析产生的标记进行比较；当我们整合两个连锁图谱的标记时，四个RIL群体中标记之间的平均遗传距离明显减小。使用这些标记进行的遗传研究成功地将与抽穗期相关的5个QTL定位到3号、6号和7号染色体上，这些QTL之前被鉴定为 、 、 、 和 。因此，GRAS-Di技术提供了一种低成本且高效的基因分型方法，以克服亲缘关系密切的日本水稻品种中狭窄的遗传多样性，并使我们能够在这种种质中生成高密度连锁图谱。
