Acta Agrestia Sinica

Acta Agrestia Sinica ›› 2018, Vol. 26 ›› Issue (6): 1473-1478.DOI: 10.11733/j.issn.1007-0435.2018.06.027

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Identification of Cytoplasmic Male Sterile Lines and Maintainer Lines of Alfalfa by SCAR Markers

Zhou Le1, WANG Ying-zhe2, Tan Jing1, XU An-kai1,2, XU Bo1   

  1. 1. College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin Province 130118, China;
    2. Agro-Biotechnology Research Institute, Jilin Academy of Agricultural Sciences, Changchun, Jilin Province 130118, China
  • Received:2018-10-29 Revised:2018-12-17 Online:2018-12-15 Published:2019-01-28

紫花苜蓿细胞质雄性不育系和保持系SCAR标记的鉴定研究

周仂1, 王英哲2, 谭晶1, 徐安凯1,2, 徐博1   

  1. 1. 吉林农业大学动物科学技术学院, 吉林 长春 130118;
    2. 吉林省农业科学院, 吉林 长春 130118
  • 通讯作者: 徐博
  • 作者简介:周仂(1994-),男,甘肃张掖人,硕士,牧草种质资源与遗传育种,E-mail:zhoule228@126.com
  • 基金资助:
    吉林省教育厅"十三五"科学技术项目（JJKH20180666KJ）；吉林省科学技术厅重点科技研发项目（20180201072ny）；国家自然科学基金青年科学基金项目（31402125）资助

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Abstract: The aim of this study was to screen molecular markers highly linked to the restoring gene of cytoplasmic male sterility in alfalfa and transform them into SCAR-specific markers. The identification and analysis showed that the marker was reproducible and stable,which provided a research basis for the production and identification of sterile line seeds in "three lines" of alfalfa. The method was to screen at the mitochondrial genome level of alfalfa with 100 ISSR primers. The UBC842 and UBS864 primers showed specific fragments between the alfalfa sterile line and its homologous maintainer line. After repeated verification,the differential bands were stable,and the band sizes were 457 bp and 645 bp,respectively. At the same time,the obtained differential bands were recovered by gel recovery method,linked to the cloning vector,sequenced and transformed into SCAR-specific identification markers. Then,two pairs of specific primers were used to identify the male sterile line and the homologous maintainer line of the newly cultured four groups of alfalfa hybrids,and the results showed that the repeatability of the SCAR marker was stable. In this study,ISSR markers were used to analyze the sterile and maintainer lines of alfalfa,and the obtained differential fragments were cloned,sequenced and transformed into SCAR-specific markers. This marker can be used as a candidate marker for seed purity analysis of alfalfa sterile lines,and is actually applied to the identification of seed purity of sterile lines.

Key words: Alfalfa, Male Sterility, SCAR Marker, Sterility Mechanism

摘要： 本研究通过已获得细胞质雄性不育系材料MS-GN-1A和保持系材料MS-GN-1B筛选与紫花苜蓿细胞质不育恢复基因高度连锁的分子标记并转化为SCAR特异标记，经鉴定分析发现该标记的重复性好，而且稳定，对紫花苜蓿“三系”配套中不育系种子生产鉴定提供研究基础。实验应用100条ISSR引物在紫花苜蓿线粒体基因组水平进行筛选，UBC842、UBS864引物在紫花苜蓿不育系及其同型保持系间显示出特异性片段，经反复验证，其差异条带均稳定出现，条带大小分别为457 bp和645 bp。对已获得的差异条带进行胶回收连接克隆载体后测序并转化为SCAR特异鉴定标记，以两对特异性引物对新培育的4组紫花苜蓿杂交种的不育系母本和同型保持系父本进行鉴定分析，发现该SCAR标记的重复性稳定。本研究通过ISSR标记对紫花苜蓿的不育系与保持系进行分析，将得到的差异片段克隆测序后转化为SCAR特异标记，该标记可以作为紫花苜蓿不育系种子纯度分析的候选标记，实际应用于不育系种子纯度的鉴定。

关键词: 紫花苜蓿, 雄性不育, SCAR标记, 不育机理

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