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甘蓝型油菜P5CR同源基因的克隆表达及其多态性分析

马秀奇 张晓娟 孙晓敏 陈乔 张羽 何开罡 权莹 宋建民

马秀奇,张晓娟,孙晓敏,等. 甘蓝型油菜P5CR同源基因的克隆表达及其多态性分析 [J]. 福建农业学报,2022,37(6):727−733 doi: 10.19303/j.issn.1008-0384.2022.06.006
引用本文: 马秀奇,张晓娟,孙晓敏,等. 甘蓝型油菜P5CR同源基因的克隆表达及其多态性分析 [J]. 福建农业学报,2022,37(6):727−733 doi: 10.19303/j.issn.1008-0384.2022.06.006
MA X Q, ZHANG X J, SUN X M, et al. Cloning, Expression, and Polymorphism of Homologous Brassica napus P5CR [J]. Fujian Journal of Agricultural Sciences,2022,37(6):727−733 doi: 10.19303/j.issn.1008-0384.2022.06.006
Citation: MA X Q, ZHANG X J, SUN X M, et al. Cloning, Expression, and Polymorphism of Homologous Brassica napus P5CR [J]. Fujian Journal of Agricultural Sciences,2022,37(6):727−733 doi: 10.19303/j.issn.1008-0384.2022.06.006

甘蓝型油菜P5CR同源基因的克隆表达及其多态性分析

doi: 10.19303/j.issn.1008-0384.2022.06.006
基金项目: 陕西省科技计划项目(2020NY-068);陕西理工大学人才启动项目(SLGRCQD2115);陕西理工大学 2020年大学生创新创业训练计划项目(S202010720022);陕西省教育厅科研项目(11JS034)
详细信息
    作者简介:

    马秀奇(2000−),男,硕士研究生,主要从事油菜分子遗传研究(E-mail:mxq20000308@163.com)

    通讯作者:

    张晓娟(1982−),女,副教授,博士,主要从事油菜分子遗传研究 (E-mail:zxj12162001@163.com

  • 中图分类号: S 562

Cloning, Expression, and Polymorphism of Homologous Brassica napus P5CR

  • 摘要:   目的  克隆油菜菌核病抗性相关基因,进一步为油菜抗病分子标记开发以及通过分子标记辅助育种途径选育抗菌核病油菜新品种提供理论基础。  方法  以高抗、高感菌核病油菜为研究材料,对甘蓝型油菜A03、C03染色体上的P5CR(Pyrroline-5-carboxylate reductase,吡咯林-5-羧酸还原酶)同源基因进行特异PCR扩增,克隆和测序及表达分析。利用DNAMAN软件对测序结果进行序列比对,寻找抗、感材料中的差异SNP位点,并分析这些位点与油菜菌核病抗性的关系。利用qPCR技术分析A03及C03染色体上P5CR同源基因在抗、感菌核病油菜材料中接种核盘菌前及接种后6 h、12 h、24 h、48 h的表达。  结果  C03染色体上的P5CR同源基因全长1457 bp,该基因在抗、感菌核病油菜材料中共有7个SNP位点,其中3个SNP位点可能与抗病性相关;A03染色体上的P5CR同源基因全长1526 bp,在抗、感菌核病油菜材料中该基因共有15个SNP位点,其中2个SNP位点可能与抗病性相关。A03及C03染色体上的P5CR基因在抗病材料接种后24 h表达量显著升高。  结论  油菜P5CR基因上存在多个可能与菌核病抗性相关的位点且在抗病材料接种后表达升高,表明P5CR可能参与油菜对菌核病的抗性反应。本研究为进一步揭示甘蓝型油菜菌核病抗病机理及油菜抗菌核病分子标记开发奠定基础。
  • 图  1  甘蓝型油菜A03及C03染色体上P5CR同源基因扩增产物电泳图

    A、B分别为A03及C03染色体上P5CR基因扩增电泳结果。Marker各条带分子量见图左侧标注。M,Marker III;1,川秦6R;2,南12R:3,中油821;4,沪16。

    Figure  1.  Electrophoretic map of amplified products of P5CR on chromosome A03 and C03 of B. napus

    A & B: Amplified electrophoretic diagram of P5CRs on chromosome A03 and C03, respectively. Molecular weight of band of marker shown on left. M: Marker III; 1: Chuanqin 6R; 2: Nan 12R; 3: Zhongyou 821; 4: Hu 16.

    图  2  甘蓝型油菜A03染色体上P5CR基因与菌核病抗性相关的多态性位点

    Figure  2.  Polymorphic sites of P5CR on A03 chromosome of B. napus related to Sclerotinia-resistance

    图  3  甘蓝型油菜C03染色体上P5CR基因多态性分析

    Figure  3.  Polymorphism of P5CR on C03 chromosome of B. napus

    图  4  qPCR分析A03及C03染色体上P5CR基因在抗、感菌核病油菜材料中的表达结果

    A、B分别为A03、C03染色体上P5CR基因的qPCR结果图。对同一时间不同材料中P5CR基因的表达量进行差异性分析(*表示P < 0.05,**表示P < 0.01)。

    Figure  4.  Expressions of P5CRs on A03 and C03 chromosomes in sclerotia-resistant and susceptible rapeseed materials detected by qPCR

    A & B: qPCR data on P5CRs on A03 and C03 chromosomes, respectively. Expressions of P5CR gene in different materials at the same time (* indicates P < 0.05, ** indicates P < 0.01).

    表  1  油菜样品的编号、品种名称和表现型

    Table  1.   Codes, variety names, and phenotypes of B. napus samples

    编号
    Number
    样本  
    Sample  
    抗性
    Resistance
    Y4 川秦6R Chuanqin 6R S
    Y7 南12R Nan 12R S
    Y8 中油821 Zhongyou 821 R
    Y12 沪16 Hu 16 R
    下载: 导出CSV

    表  2  甘蓝型油菜C03及A03染色体上P5CR同源基因引物序列信息

    Table  2.   Primer sequence of P5CR on chromosomes C03 and A03 of B. napus

    引物名称
    Primer name
    序列(5′-3′)
    Sequence(5′-3′)
    产物大小
    Product length/bp
    P5CRC03F2 GCCTTGGTAAGCGAATGG 1879
    P5CRC03R2 TCGTACCCTGTGACGATTCA
    P5CRA03F1 TGTGTTGGGCCTTTGTAAAACAAT 1977
    P5CRA03R1 TGCAATTTGGTCGTACCCTTTAAC
    qP5CR A03 F1 GGAAGTGGACCAGCATACG 197
    qP5CR A03 R1 GCTATTGTAGTCCCGCCA
    qP5CR C03 F1 TCGGAACAGCGGCAAGTGA 257
    qP5CR C03R1 CTTCCCTGTCTTGCTCACCAT
    Ubc21F2 TCCCGAACCGTATCCTCTGC 156
    Ubc21R2 GGTTACCTGAGTCGCAGTTGAG
    下载: 导出CSV
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出版历程
  • 收稿日期:  2022-03-08
  • 修回日期:  2022-04-27
  • 网络出版日期:  2022-06-20
  • 刊出日期:  2022-06-28

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