Hu Wei, Zhou Tianhao, Hu Gang, Wu Hong, Han Zhongmin, Xiao Jinghua, Li Xianghua, Xing Yongzhong
National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan, 430070, China.
Hubei Collaborative Innovation Center for Grain Industry, Yangtze University, Jingzhou, 434025, China.
Plant J. 2020 Nov;104(4):1129-1141. doi: 10.1111/tpj.14969. Epub 2020 Sep 2.
Spontaneous mutants are mainly obtained from tissue culture or natural occurrences in plants. The traditional strategy for identifying spontaneously mutated genes is to continuously backcross these mutants to another variety and develop a near-isogenic F population for map-based cloning or bulked segregant analysis. However, this strategy is time-consuming. Here, we have developed a new method to efficiently accelerate the identification process. The chemical mutagen ethyl methanesulfonate was first used to treat the wild type of the spontaneous mutants to induce thousands of neutral mutations. An induced individual without any statistically significant phenotypic changes which was compared with the wild type was chosen as the neutral mutant. The spontaneous mutant was then crossed with the neutral mutant to develop a pseudo-near-isogenic F population in which only the induced neutral mutations and the causal mutation were segregated in the genome. This population ensures that the variation of the mutated trait is controlled only by the spontaneously mutated gene. Finally, after sequencing the neutral mutant and the mutant-type DNA pool of the F population the spontaneous mutation will be identified quickly by bioinformatics analysis. Using this method, two spontaneously mutated genes were identified successfully. Therefore, the neutral mutant-bridging method efficiently identifies spontaneously mutated genes in rice, and its value in other plants is discussed.
自发突变体主要从植物的组织培养或自然发生中获得。鉴定自发突变基因的传统策略是将这些突变体与另一个品种连续回交，并为基于图谱的克隆或混合分离分析构建近等基因F群体。然而，这种策略耗时较长。在此，我们开发了一种新方法来有效加速鉴定过程。首先使用化学诱变剂甲基磺酸乙酯处理自发突变体的野生型，以诱导数千个中性突变。选择一个与野生型相比无任何统计学显著表型变化的诱导个体作为中性突变体。然后将自发突变体与中性突变体杂交，构建一个假近等基因F群体，其中基因组中仅分离出诱导的中性突变和致病突变。这个群体确保了突变性状的变异仅由自发突变基因控制。最后，对中性突变体和F群体的突变型DNA池进行测序后，通过生物信息学分析可快速鉴定出自发突变。利用这种方法，成功鉴定出了两个自发突变基因。因此，中性突变体搭桥法能有效鉴定水稻中的自发突变基因，并讨论了其在其他植物中的价值。
