棉花细胞质雄性不育恢复基因Rf1候选区域SSCP标记开发与候选基因表达分析

doi:10.11963/1002-7807.llcxcz.20180104

摘要/Abstract

摘要： 【目的】棉花细胞质雄性不育恢复基因Rf1的定位对于研究恢复基因的作用机制和改良恢复系具有重要意义。前期研究结果表明在恢复基因Rf₁所在Scaffold 333上的目标区间内有9个PPR（Pentatricopeptide repeat）基因成簇分布在160 kb区间内，此区间内还包括另外9个其他类型的基因。为进一步丰富恢复基因目标区域内标记数目，并了解相关基因的表达模式。【方法】利用这18个基因的序列设计出覆盖全部基因的引物共155对，通过单链构象多态性（Single-strand conformation polymorphism，SSCP）技术对可育池和不育池进行筛选，利用荧光定量聚合酶链式反应（Polymerase chain reaction，PCR）对目标区域内的8个非PPR基因在不育系、保持系、恢复系的花蕾不同发育时期的表达模式进行了分析。【结果】共得到15对多态性引物，结合前期的3个简单重复序列标记（Simple sequence repeats，SSR）标记对F₂群体进行基因型分析，结果表明这些标记分布在4.8 cM范围内。受恢复基因或不育胞质的影响，在花蕾4个发育时期，大部分基因表达存在明显差异。【结论】本研究获得了与恢复基因紧密连锁的SSCP标记，并初步了解了目标区间内部分基因的表达模式，为进一步开展恢复基因精细定位和候选基因的筛选提供了基础。

关键词: 棉花; 恢复基因Rf1; 遗传定位; PPR基因; SSCP标记

Abstract: [Objective] Locating the cotton cytoplasmic male sterility (CMS) restorer gene Rf₁ is important for investigating restorer gene mechanisms and improving restorer lines. In our previous study, a gene cluster, with nine Pentatricopeptide repeat(PPR) genes and nine other genes, was found within the 160-kb Rf₁target region in Scaffold 333. The objective here was to improve the density of Rf1-linked markers in the target region and determine the expression profiles of candidate genes. [Method] Using the sequences of the 18 genes, we designed 155 single-strand conformation polymorphism (SSCP) primers covering all of the gene sequences to identify the polymorphic SSCP markers between the fertile and sterile pools. Additionally, real-time polymerase chain reaction(PCR) was performed to analyze the expression profiles of eight candidate genes in the four developmental stages of buds of sterile, maintainer and restoring lines, respectively. [Result] In total, 15 polymorphic primers were identified. A genotype analysis of the F₂ population was conducted using the 15 primers and 3 other polymorphic simple sequence repeats (SSR) markers. The markers were distributed in a 4.8 cM range. In addition, owing to the influence of sterile cytoplasm or restorer genes, most of the genes showed different expression patterns in the four developmental stages of the three lines' buds. [Conclusion] SSCP markers tightly linked to Rf₁ were identified and the expression profiles of candidate genes were determined. This study provides a basis for the further fine mapping of restorer genes and for candidate gene screening.

Key words: cotton; restorer genes; genetic mapping; PPR gene; SSCP marker

中图分类号:

S562.03：S321.8

刘利彩, 郭立平, 戚廷香, 王海林, 唐会妮, 张学贤, 乔秀琴, 吴建勇, 邢朝柱. 棉花细胞质雄性不育恢复基因Rf₁候选区域SSCP标记开发与候选基因表达分析[J]. 棉花学报, 2018, 30(1): 12-20.

Liu Licai, Guo Liping, Qi Tingxiang, Wang Hailin, Tang Huini, Zhang Xuexian, Qiao Xiuqin, Wu Jianyong, Xing Chaozhu. SSCP Marker Development and Analysis of Candidate Restorer Gene Rf₁for Cotton Cytoplasmic Male Sterility[J]. Cotton Science, 2018, 30(1): 12-20.

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棉花细胞质雄性不育恢复基因Rf₁候选区域SSCP标记开发与候选基因表达分析

SSCP Marker Development and Analysis of Candidate Restorer Gene Rf₁for Cotton Cytoplasmic Male Sterility

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