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作物学报 ›› 2023, Vol. 49 ›› Issue (10): 2603-2612.doi: 10.3724/SP.J.1006.2023.21082

• 作物遗传育种·种质资源·分子遗传学 •    下一篇

Pm21基因的次级易位创制及鉴定

张蓝月1(), 罗江陶2(), 范超兰1, 李亚洲1, 姜博1, 陈雪1, 陈雪姣1, 袁中伟1, 甯顺腙1, 张连全3, 刘登才3(), 郝明1()   

  1. 1四川农业大学小麦研究所, 四川温江 611130
    2四川省农业科学院作物研究所, 四川锦江 610066
    3西南作物基因资源发掘与利用国家重点实验室, 四川温江 611130
  • 收稿日期:2022-12-12 接受日期:2023-02-21 出版日期:2023-10-12 网络出版日期:2023-03-06
  • 通讯作者: 郝明, E-mail: haomingluo@foxmail.com; 刘登才, E-mail: dcliu7@sicau.edu.cn
  • 作者简介:张蓝月, E-mail: 569395258@qq.com;罗江陶, E-mail: jtluohao@163.com**同等贡献
  • 基金资助:
    四川省科学技术项目(2022ZDZX0014);四川省科学技术项目(2022NSFSC1696);国家自然科学基金项目(31971884);国家自然科学基金项目(32172020)

Creation and analysis of secondary translocation harbouring gene Pm21

ZHANG Lan-Yue1(), LUO Jiang-Tao2(), FAN Chao-Lan1, LI Ya-Zhou1, JIANG Bo1, CHEN Xue1, CHEN Xue-Jiao1, YUAN Zhong-Wei1, NING Shun-Zong1, ZHANG Lian-Quan3, LIU Deng-Cai3(), HAO Ming1()   

  1. 1Triticeae Research Institute, Sichuan Agricultural University, Wenjiang 611130, Sichuan, China
    2Crop Research Institute, Sichuan Academy of Agricultural Science, Jinjiang 610066, Sichuan, China
    3State Key Laboratory of Crop Gene Exploration and Utilization in Southwest China, Sichuan Agricultural University, Wenjiang 611130, Sichuan, China
  • Received:2022-12-12 Accepted:2023-02-21 Published:2023-10-12 Published online:2023-03-06
  • Contact: E-mail: haomingluo@foxmail.com; E-mail: dcliu7@sicau.edu.cn
  • About author:**Contributed equally to this study
  • Supported by:
    Sichuan Science and Technology Program(2022ZDZX0014);Sichuan Science and Technology Program(2022NSFSC1696);National Natural Science Foundation of China(31971884);National Natural Science Foundation of China(32172020)

摘要:

小麦-簇毛麦6VS.6AL易位携带抗白粉病基因Pm21, 对我国小麦抗白粉病育种做出了重要贡献。本文对162份四川小麦品种(系)的55K SNP芯片数据分析表明, 25份含6VS.6AL易位染色体, 占15.4%。重组位置和单倍型分析表明, 这些材料的6VS.6AL均为着丝点易位、具有单一来源。根据系谱分析, 92R178为最原始的供体材料。利用由ph1b诱导6VS/6AS部分同源重组形成的, 含Pm21基因的初级易位6VS-6AS.6AL和6AS-6VS.6AL为亲本, 创制了1个含Pm21基因、6VS片段大幅减小的6AS-6VS-6AS.6AL次级易位。根据中国春参考基因组, 该次级易位的2个重组位点, 分别位于6A染色体53.1~53.8 Mb和90.7~92.2 Mb之间, 易位片段大小在36.9~39.1 Mb之间。分子细胞学鉴定表明, ph1b诱导外源易位的同时, 小麦自身的内源染色体也发生了大量易位, 这影响易位系的遗传稳定和育种利用。为了解决这个问题, 建议将ph1b基因突变系以杂合的形式进行长期保存。同时, 在利用ph1b诱导小麦-外源易位的过程中, 尽量减少ph1b纯合状态下的繁殖世代, 以减少小麦内源染色体易位。育种利用时, 应尽快消除小麦内源易位。

关键词: 小麦, 簇毛麦, 白粉病, 6VS.6AL易位, Pm21基因, 小片段易位系

Abstract:

Wheat-Haynaldia villosa 6VS.6AL translocation harbouring the gene Pm21 has made a great contribution to powdery mildew resistance breeding in China. Based on the data of 55K SNP chip, 25 (15.4%) out of 162 Sichuan wheat varieties contained the translocation. In this study, recombination point and haplotype analysis on the 25 varieties showed that it was centric translocation. Combined with the pedigree information, 92R178 was the original donor of the 6VS.6AL translocation in these varieties. 6AS-6VS-6AS.6AL secondary recombinant containing Pm21 was generated by using primary recombinants 6VS-6AS.6AL and 6AS-6VS.6AL as the cross parents, which both formed by the induction of ph1b. The secondary recombinant had a much smaller 6VS chromatin than the primary recombinants. Based on the Chinese Spring reference genome, the crossover points of the secondary recombinant were located within 53.1-53.8 Mb and 90.7-92.2 Mb of chromosome 6A, with a 6VS fragment size about 36.9-39.1 Mb. Molecular cytological identification also detected the extensive recombinants among wheat endogenous homoeologs induced by ph1b, which was not only disadvantage for genetic stabilization of wheat-alien recombinants but also for wheat breeding. A proposed solution to reduce endogenous recombinants was to preserve the ph1b mutant line in a heterozygous condition and reduce the selfing times during the development of ph1b-mediated wheat-alien recombination. In breeding, it is necessary to eliminate endogenous recombinants as soon as possible.

Key words: wheat, Haynaldia villosa, powdery mildew, 6VS.6AL translocation, Pm21 gene, small fragment translocation line

表1

25份携带6VS.6AL易位的四川小麦品种(系)"

品种(系)名
Accession name
选育单位
Origin
杂交组合
Pedigree
审定年份
Released year
细胞学*
Cytological*
绵阳27
Mianyang 27
绵阳市农业科学院
Mianyang Academy of Agricultural Sciences
81-5/81-24 1997 /
绵阳28
Mianyang 28
绵阳市农业科学院
Mianyang Academy of Agricultural Sciences
T79350-1-4/Mianyang 11 1997 /
川麦36
Chuanmai 36
四川省农业科学院
Sichuan Academy of Agricultural Sciences
Milan/SW5193 2002 ×
内麦8号
Neimai 8
内江市农业科学院
Neijiang Academy of Agricultural Sciences
Mianyang 26/92R178 2003
西科麦1号
Xikemai 1
西南科技大学
SouthWest University of Science and Technology
Mianyang 88-304/Mo-212 2003 ×
良麦2号
Liangmai 2
四川农业大学
Sichuan Agricultural University
Mianyang 26/// (10-A/88-1643//Chuanyu 12) 2004 ×
绵麦37
Mianmai 37
绵阳市农业科学院
Mianyang Academy of Agricultural Sciences
96EW37/Mianyang 90-100 2004
内麦9号
Neimai 9
内江市农业科学院
Neijiang Academy of Agricultural Sciences
Mianyang 26/92R178 2004
杏麦2号
Xingmai 2
内江市农业科学院
Neijiang Academy of Agricultural Sciences
Mianyang 26/92R178 2004
内麦11号
Neimai 11
内江市农业科学院
Neijiang Academy of Agricultural Sciences
Mianyang 26/92R178 2007
绵麦185
Mianmai 185
绵阳市农业科学院
Mianyang Academy of Agricultural Sciences
Mianyang 96-5/Liaochun 10 2008
内麦3416
Neimai 3416
内江市农业科学院
Neijiang Academy of Agricultural Sciences
R57/Ping 5 2010 /
绵麦228
Mianmai 228
绵阳市农业科学院
Mianyang Academy of Agricultural Sciences
1275-1/Nei 2938//99-1522 2011 ×
绵麦51
Mianmai 51
绵阳市农业科学院
Mianyang Academy of Agricultural Sciences
1275-1/99-1522 2012
西科麦7号
Xikemai 7
西南科技大学
Southwest University of Science and Technology
Chuanyu 11/Mo444 2012
Y11-1741 四川农业大学
Sichuan Agricultural University
Zimai 1/Mianyang 2003-1 2012
绵麦1618
Mianmai 1618
绵阳市农业科学院
Mianyang Academy of Agricultural Sciences
1275-1//Nei 2938/99-1522 2013
西科麦8号
Xikemai 8
西南科技大学
Southwest University of Science and Technology
97-392/Yun 225747-5 2013
西科麦9号
Xikemai 9
西南科技大学
Southwest University of Science and Technology
Nei 4301/Mianyang 31 2014
宜麦9号
Yimai 9
宜宾市农业科学院
Yibin Academy of Agricultural Sciences
R59/Yi 97-24 2014 ×
川麦92
Chuanmai 92
四川省农业科学院
Sichuan Academy of Agricultural Sciences
Neimai 8/Jian 3//Chuanmai 42 2015 /
绵麦112
Mianmai 112
绵阳市农业科学院
Mianyang Academy of Agricultural Sciences
Mian 06-367/99-1522 2015 /
内麦366
Neimai 366
内江市农业科学院
Neijiang Academy of Agricultural Sciences
Balandal88/Nei 4344 2015 /
国豪麦3号
Guohaomai 3
四川国豪种业
Sichuan Guohao Seeds Industry Co., Ltd.
1227-185/99-1522//99-1572 2016
绵麦285
Mianmai 285
绵阳市农业科学院
Mianyang Academy of Agricultural Sciences
1275-1/99-1522 2016

图1

162份四川小麦6A (6VS.6AL)染色体基因型 图中每一行代表1个品种(系); 每一列代表1个标记, 标记按照IWGSC v1.0参考基因组6A短臂至长臂的顺序从左向右排列。每个标记的分型结果分别按照A (红色)、C (绿色)、G (黄色)、T (蓝色)、杂合H (灰色)、缺失NA (粉色)表示。Hap1~3代表近着丝粒无重组区划分的单倍型, 括号里面数字代表归属该单倍型的品种(系)数。箭头示在IWGSC v1.0参考基因组上的位置。"

图2

6VS/6AS次级易位创制流程及细胞学鉴定 A: 利用6VS/6AS易位创制次级易位的杂交和筛选流程; B: Pm21特异标记CINAU-NLR1标记鉴定(从左至右, 依次为marker、Pm99915-1、CSph1b、HM782-15和HM780-7); C: 次级易位系Rec32的基因组原位杂交鉴定结果; D: 次级易位系Rec50的基因组原位杂交鉴定结果; E: 6VS/6AS初级易位重组重新形成6VS.6AL易位系的基因组原位杂交鉴定结果; F: 初级易位配对重组形成次级易位示意图。MI: 减数分裂中期I; AI: 减数分裂后期I。6VS染色体臂或片段在模式染色体中用红色填充表示, 6A染色体或片段用蓝色填充表示, 黄色短横线则代表Pm21基因。黄色箭头指示经同源重组形成的次级易位C_6VS/6AS_AVA-Pm21-1或6VS.6AL臂间易位, 白色箭头指示未参与重组的初级易位。"

图3

Rec50和HM887-9-51的16K缺失标记分布图 图中每个染色体的左边表示Rec50, 右边表示HM887-9-51。缺失标记位置用绿色表示, 非缺失标记用红色表示。实线框指示非目标小麦内源结构变异染色体区段, 虚线框指示目标次级和初级易位缺失6AS片段。"

图4

Rec50染色体组成鉴定 a: 基因组原位杂交鉴定。A基因组标记为粉红色, B基因组标记为蓝色, D基因组标记为绿色。b: 荧光原位杂交鉴定。探针Oligo-pSc119.2标记为绿色, Oligo-pTa535标记为粉红色。白色箭头指示6VS/6AS目标染色体臂以外的小麦染色体间重组, 红色箭头指示6VS/6AS目标初级和次级重组染色体。"

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