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农学学报

农学学报 ›› 2023, Vol. 13 ›› Issue (12): 20-27.doi: 10.11923/j.issn.2095-4050.cjas2022-0153

• 农艺科学 生理生化 • 上一篇    下一篇

抗根肿病大白菜材料SSR分子标记的初步筛选

兰梅1,2(), 张丽琴1,2, 徐学忠1,2, 胡靖锋1,2, 杨红丽1,2, 杨鼎3, 李崇娟3, 和江明1,2()   

  1. 1 云南省农业科学院园艺作物研究所，昆明 650205
    2 国家蔬菜改良分中心云南分中心，昆明 650205
    3 云南大学，昆明 650504
  • 收稿日期:2022-11-09 修回日期:2023-05-15 出版日期:2023-12-20 发布日期:2023-12-19
  • 通讯作者:
    和江明，男，1969年出生，云南香格里拉人，研究员，本科，主要从事十字花科蔬菜育种与栽培研究。通信地址：650205 云南省昆明市盘龙区北京路2238号园艺所，E-mail：。
  • 作者简介:

    兰梅，女，1986年出生，云南昆明人，助理研究员，硕士，研究方向：十字花科蔬菜分子生物学辅助育种。通信地址：650205 云南省昆明市盘龙区北京路2238号园艺所，E-mail：。

  • 基金资助:
    财政部和农业农村部“国家现代农业产业技术体系”(CARS-23-G-33); 云南省科技厅科技计划项目基础研究专项“青花菜花球遗传图谱的构建及硫代葡萄糖苷相关性状的QTL定位分析”(202301AT070089); 云南省发展和改革委员会“万人计划产业领军人才”(YNWR-CYJS-2018-040); 云南省科技厅科技计划项目“通海县蔬菜产业科技特派团”(202204BI090023); 云南省科技厅重大科技专项计划子课题“云南野、特蔬菜种质保护及研究利用”(202302AE090006-4)

Preliminary Screening of SSR Molecular Markers for Resistance to Clubroot in Brassica rapa L. ssp. pekinensis

LAN Mei1,2(), ZHANG Liqin1,2, XU Xuezhong1,2, HU Jingfeng1,2, YANG Hongli1,2, YANG Ding3, LI Chongjuan3, HE Jiangming1,2()   

  1. 1 Horticultural Crops Research Institute, Yunnan Academy of Agricultural Sciences, Kunming 650205, Yunnan, China
    2 YunnanBranch of National Vegetable Improvement Center, Kunming 650205, Yunnan, China
    3 Yunnan University, Kunming 650504, Yunnan, China
  • Received:2022-11-09 Revised:2023-05-15 Online:2023-12-20 Published:2023-12-19

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摘要：

为了明确抗根肿病大白菜材料CCR12049中的抗病基因，进一步开发分子标记。以高抗根肿病的高代自交系大白菜CCR12049、高感根肿病的高代自交系大白菜CM12081、CCR12049和CM12081杂交得到的F1及F1自交构建的F2分离群体为试材，通过人工接种鉴定、聚丙烯酰胺凝胶电泳和序列比对。结果显示，该抗病材料中的根肿病抗性由显性单基因控制；36对引物在2个亲本和F1中初步筛选出4对有多态性的引物，在F2中进一步验证，发现只有1对(cr-26)在F2中的扩增结果与表型鉴定一致；2个亲本及F1的PCR产物测序比对发现，在95~111 bp这个位置，抗病亲本和F1的序列完全相同，但是感病材料在这个位置出现了17个碱基(TCTCTATCTCTTACGCA)的缺失。可以推断抗病材料可能是由于这17个碱基的插入从而表现出抗病性，该标记可以将抗感材料区分开，该标记可以作为一个初步筛选白菜抗根肿病的SSR分子标记来利用。

关键词: 大白菜, 根肿病, SSR分子标记, 遗传规律, 序列比对

Abstract:

In order to identify the clubroot resistance gene in Chinese cabbage cultivar CCR12049 and develop molecular markers, the following materials were employed: CCR12049, a high-generation inbred line with high resistance to clubroot disease, CM12081, a high-generation inbred line with high susceptibility to clubroot disease, F1 generation resulting from the cross between CCR12049 and CM12081, and the F2 generation segregating populations created through selfing of the F1 generation. Through artificial inoculation identification, polyacrylamide gel electrophoresis, and sequence alignment, it was demonstrated that the disease resistance in tested materials was governed by a dominant single gene. 36 primer pairs were initially tested on both the parental lines and F1 generation, 4 primer pairs exhibiting polymorphism were identified. Subsequent validation in the F2 generation revealed that only one pair (cr-26) yielded amplification results consistent with the observed phenotype in the F2 generation. Through sequencing comparison of the PCR products obtained from the two parental lines and F1, it was observed that the sequences of the resistant parents and F1 were identical within the range of 95 bp to 111 bp. Conversely, in the susceptible materials, a deletion of 17 bases (TCTCTATCTCTTACGCA) was identified. These findings suggest that the resistance observed in the impermeable materials may be attributed to the insertion of these 17 bases. This marker has the capability to distinguish the resistant materials and can serve as an SSR marker for the initial screening of clubroot resistance in Chinese cabbage.

Key words: Chinese cabbage, clubroot disease, simple sequence repeats (SSR) molecular marker, inheritance law, sequence alignment