Koshika Natsume, Shioya Naohiro, Fujimura Takashi, Oguchi Rina, Ota Chie, Kato Emi, Takahashi Reiko, Kimura Shuichi, Furuno Shinsuke, Saito Koichi, Okabe Kazuhiro, Watanabe Masanori, Hoshino Tomoki
Laboratory of Crop Breeding, Graduate School of Agricultural Sciences, Yamagata University, Wakaba-machi 1-23, Tsuruoka 997-8555, Yamagata, Japan.
Laboratory of Crop Breeding, Faculty of Agriculture, Yamagata University, Wakaba-machi 1-23, Tsuruoka 997-8555, Yamagata, Japan.
Plants (Basel). 2022 Jul 13;11(14):1839. doi: 10.3390/plants11141839.
Induced mutation is a viable breeding strategy that is widely utilized in the development of elite plant varieties. We aimed to improve a variety of edamame by constructing novel mutant populations using the ethyl methanesulfonate in soybeans ( (L.) Merr.). In the M population, the flowering stage showed a considerable standard deviation compared to the wild type, confirming that the mutant populations had the expected DNA mutations. To identify the DNA mutations in the mutant populations, we used the targeting induced local lesions in genomes (TILLING) method, which is a reverse genetic method, to search for soybean flowering-related gene mutants. A total of 30 mutants from , , , and genes, which are known to be highly effective genes, or their homologous gene for flowering and maturation found in soybean quantitative trait locus analyses were isolated from our TILLING screening. Among these mutants, there were eleven nonsynonymous substitution mutants, one nonsense mutant, and two single nucleotide deletion mutants that could be expected to reduce or eliminate gene function. The , , and mutants obtained in this study flowered considerably earlier than the wild type. In particular, the mutant with a nonsynonymous substitution flowered approximately 1 month after sowing regardless of the sowing date, and its harvest date was approximately 1 month earlier than that of the wild type. Mutations identified using the TILLING method could not only be used as gel-based DNA markers with the same manipulation method, but the mutations could also be detected as DNA markers by the high-resolution melting method. These results indicate that mutations achieved without chromosome modification by crossbreeding are effective for early and practical improvement of superior varieties and that efficient selection of mutants by reverse genetics is an effective method for the identification of genetic modifications. The edamame mutant populations developed in this study are believed to possess various useful alleles which may be applicable in the search for mutations that lead to improved edamame yield and eating quality beyond the flowering stage.
诱变是一种可行的育种策略，在优良植物品种的培育中被广泛应用。我们旨在通过用甲基磺酸乙酯处理大豆（(L.) Merr.）构建新的突变群体来改良毛豆品种。在M群体中，与野生型相比，开花期表现出相当大的标准差，证实突变群体具有预期的DNA突变。为了鉴定突变群体中的DNA突变，我们使用了基因组靶向诱导局部损伤（TILLING）方法，这是一种反向遗传学方法，来寻找大豆开花相关基因突变体。从我们的TILLING筛选中分离出了总共30个来自大豆数量性状位点分析中已知的高效基因、开花和成熟相关基因或其同源基因的突变体。在这些突变体中，可以预期有11个非同义替换突变体、1个无义突变体和2个单核苷酸缺失突变体能够降低或消除基因功能。本研究中获得的、和突变体开花时间比野生型早得多。特别是，具有非同义替换的突变体无论播种日期如何，播种后约1个月开花，其收获日期比野生型早约1个月。使用TILLING方法鉴定的突变不仅可以作为凝胶基DNA标记以相同的操作方法使用，而且这些突变也可以通过高分辨率熔解方法作为DNA标记进行检测。这些结果表明，通过杂交在不进行染色体修饰的情况下实现的突变对于优良品种的早期和实际改良是有效的，并且通过反向遗传学对突变体进行高效选择是鉴定遗传修饰的有效方法。本研究中开发的毛豆突变群体被认为拥有各种有用的等位基因，这些等位基因可能适用于寻找导致毛豆产量和食用品质在开花期之后得到改善的突变。
