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作物学报 ›› 2020, Vol. 46 ›› Issue (7): 997-1005.doi: 10.3724/SP.J.1006.2020.94143

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

大豆品种郑97196抗疫霉病基因RpsZheng精细定位

张雪翠1,钟超2,段灿星1,孙素丽1,*(),朱振东1,*()   

  1. 1 中国农业科学院作物科学研究所, 北京100081
    2 沈阳农业大学农学院, 辽宁沈阳110866
  • 收稿日期:2019-09-27 接受日期:2020-01-15 出版日期:2020-07-12 网络出版日期:2020-02-22
  • 通讯作者: 孙素丽,朱振东
  • 作者简介:E-mail: 553870151@qq.com
  • 基金资助:
    国家公益性行业(农业)科研专项经费项目(201303018);农业农村部农作物种质资源保护与利用专项(2019NWB036-12);中国农业科学院科技创新工程项目

Fine mapping of Phytophthora resistance gene RpsZheng in soybean cultivar Zheng 97196

ZHANG Xue-Cui1,ZHONG Chao2,DUAN Can-Xing1,SUN Su-Li1,*(),ZHU Zhen-Dong1,*()   

  1. 1 Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing 100081, China
    2 Agricultural College, Shenyang Agricultural University, Shenyang 110866, Liaoning, China
  • Received:2019-09-27 Accepted:2020-01-15 Published:2020-07-12 Published online:2020-02-22
  • Contact: Su-Li SUN,Zhen-Dong ZHU
  • Supported by:
    Special Fund for Agroscientific Research in the Public Interest(201303018);Program of Protection of Crop Germplasm Resources from the Ministry of Agriculture and Rural Affairs(2019NWB036-12);Scientific Innovation Program of Chinese Academy of Agricultural Sciences

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摘要：

大豆疫霉病是由大豆疫霉引起的一种重要大豆病害, 可造成严重的经济损失。种植含有抗疫霉病基因的大豆品种是控制该病害最有效的途径。前人在大豆品种郑97196的3号染色体上鉴定了一个抗疫霉病基因RpsZheng。本研究的目的是验证并精细定位抗疫霉病基因RpsZheng。以Williams和郑97196杂交衍生的188个F2:3家系为作图群体, 用大豆3号染色体上的SSR标记构建RpsZheng遗传连锁图, 获得与RpsZheng紧密连锁的侧翼SSR标记SattWM82_39 (2.5 cM)和BARCSOYSSR_03_0269 (1.0 cM)。基于亲本间全基因组重测序数据鉴定和开发多态性InDel标记, 进一步将RpsZheng候选区域缩小至105.2 kb, 通过检测RpsZheng候选区域内的共分离标记特异性, 获得了能够有效检测RpsZheng的分子标记WZInDel11。本研究明确了RpsZheng的候选基因组区间, 鉴定出了能够有效用于基因功能研究和辅助选择育种的共分离分子标记。

关键词: 大豆, 大豆疫霉病, 抗性遗传分析, 抗病基因, 精细定位

Abstract:

Phytophthora root rot caused by Phytophthora sojae, is a major disease in soybean production, which can cause serious yield losses. To date, the most effective way to control the disease is deployment of resistant soybean cultivars containing Phytophthora sojae (Rps) resistance genes. The previous study identified an Rps gene RpsZheng in soybean cultivar Zheng 97196, which was mapped on chromosome 3. The objective of this study was to confirm and finely map the resistance gene RpsZheng. The susceptible Williams and resistant Zheng 97196 were crossed and generated 188 F2:3 families, which were used as a mapping population. The genetic linkage map of RpsZheng was constructed using genetic and phenotypic data. RpsZheng was mapped between the flanking SSR markers SattWM82_39 (2.5 cM) and BARCSOYSSR_03_0269 (1.0 cM). Based on genome-wide resequencing data of parents, we developed and identified polymorphic InDel markers to further narrow the RpsZheng candidate region to 105.2 kb. Molecular detection among soybean cultivars was carried out using the co-segregation markers in the RpsZheng candidate region. The marker WZInDel11 could effectively distinguish RpsZheng from other Rps genes. This study identified specific genomic intervals of RpsZheng and developed co-segregation markers that can be effectively used for gene function studies and molecular assisted selection breeding.

Key words: Glycine max, Phytophthora root rot, genetic analysis of resistance, resistance gene, fine mapping

表1

28个大豆品种对10个大豆疫霉分离物的抗性反应型"

品种(抗疫霉病基因)
Cultivar (Rps gene)		 大豆疫霉分离物Phytophthora sojae isolate
PsRace1 PsRace4 PsJS2 PsNKI Ps41-1 PsMC1 PsAH4 Ps7063 Ps6497 PsUSAR2
Harlon (Rps1a) S S S R S S R S R R
Harosoy 13XX (Rps1b) R R S S S S S R S S
Williams 79 (Rps1c) R R S R R R S R R R
PI103091 (Rps1d) R S S S S S S S S R
Williams 82 (Rps1k) R R S R R S S R R R
L76-988 (Rps2) R R S S S S S S S S
L83-570 (Rps3a) R R S S S S S S R R
PRX146-36 (Rps3b) R S S S S S S R R R
PRX145-48 (Rps3c) R R S S S S S R S S
L85-2352 (Rps4) R R S S S S S S R R
L85-3059 (Rps5) R R S S S S S S R S
Harosoy 62XX (Rps6) R R S S S S S S R R
Harosoy (Rps7) R R S S S S S S S R
PI399073 (Rps8) R R S S S S R S R R
鲁豆4号 Ludou 4 (Rps9) R R R R R R R R R R
皖豆15 Wandou 15 (Rps10) R R S R R R R S R R
诱变30 Youbian 30 (RpsYB30) R S S R R S S S S R
豫豆29 Yudou 29 (RpsYD29) R R S R R R S R R R
齐茶豆1号 Qichadou 1 (RpsQ) R R R R R R R R R R
华春18 Huachun 18 (RpsHC18) R R R R R R R S R R
豫豆25 Yudou 25 (RpsYD25) R R S R R R S R R R
早熟18 Zaoshu 18 (RpsZS18) R R S R R S S S R R
岫94-11 Xiu 94-11 (RpsX) R R R R R R R R R R
郑97196 Zheng 97196 (RpsZheng) R R R R R R R R R R
中黄13 Zhonghuang 13 (rps) S S S S S S S S S S
豫豆21 Yudou 21 (rps) S S S S S S S S S S
Williams (rps) S S S S S S S S S S

附表1

用于验证共分离分子标记的226个已知抗性表型的大豆品种"

对大豆疫霉分离物PsJS2的抗性反应 Phenotypic reaction to PsJS2
不含有WZInDel11目标片段的大豆品种
Soybean cultivar without WZInDel11 target fragment		 豫豆1号(S)、豫豆2号(S)、豫豆3号(S)、豫豆6号(S)、豫豆16号(S)、豫豆18号(S)、豫豆20号(S)、豫豆21号(S)、豫豆22号(S)、豫豆26号(S)、豫豆27号(S)、郑120(S)、郑77249(S)、郑85558(S)、郑90007(S)、周豆17(S)、周豆18(S)、中黄1号(S)、中黄2号(S)、中黄3号(S)、中黄4号(S)、中黄5号(S)、中黄6号(S)、中黄7号(S)、中黄8号(S)、中黄9号(S)、中黄10号(S)、中黄11号(S)、中黄12号(S)、中黄14号(S)、中黄15号(S)、中黄16号(S)、中黄17号(S)、中黄18号(S)、中黄19号(S)、中黄20号(S)、中黄21号(S)、中黄22号(S)、中黄23号(S)、中黄24号(S)、中黄25号(S)、中黄26号(S)、中黄27号(S)、中黄28号(S)、中黄29号(S)、中黄30号(S)、中黄31号(S)、中黄33号(S)、中黄34号(S)、中黄35号(S)、中黄36号(S)、中黄37号(S)、中黄38号(S)、中黄39号(S)、中黄40号(S)、中黄41号(S)、中黄42号(S)、中黄43号(S)、中黄44号(S)、中黄45号(S)、中黄46号(S)、中黄47号(S)、中黄48号(S)、中黄49号(S)、中黄50号(S)、中黄52号(S)、中黄53号(S)、中黄55号(S)、中黄56号(S)、中黄57号(S)、中黄58号(S)、中黄59号(R)、中黄61号(S)、中黄62号(S)、中黄63号(S)、中黄64号(S)、中黄66号(S)、中黄68号(S)、中黄69号(S)、中黄70号(S)、中黄71号(S)、中黄74号(S)、菏豆1号(S)、菏豆2号(S)、菏豆13号(S)、菏豆14号(S)、菏豆15号(S)、菏豆16号(S)、菏豆17号(S)、菏豆18号(S)、菏豆19号(S)、菏豆20号(S)、菏豆21号(S)、菏豆22号(S)、菏豆23号(S)、菏豆24号(S)、菏豆25号(S)、菏豆26号(S)、菏豆27号(S)、淮豆1号(S)、淮豆2号(S)、淮豆3号(S)、淮豆4号(S)、淮豆5号(S)、淮豆6号(S)、淮豆7号(S)、淮豆11号(S)、鲁0110-1(S)、鲁0110-8(S)、鲁0126(S)、鲁0413(R)、鲁97011-1-11(S)、鲁99013-30-1(S)、鲁豆1(S)、鲁豆2(S)、鲁豆3(S)、鲁豆7(S)、齐黄2号(S)、齐黄5号(S)、齐黄6号(S)、齐黄7号(S)、齐黄8号(S)、齐黄9号(S)、齐黄10号(S)、齐黄12号(S)、齐黄13号(S)、齐黄20号(S)、齐黄21号(S)、皖豆1号(S)、皖豆2号(S)、皖豆3号(S)、皖豆5号(S)、皖豆6号(S)、皖豆7号(S)、皖豆9号(S)、皖豆12(S)、皖豆14(S)、皖豆16(S)、皖豆17(S)、皖豆18(S)、皖豆19(S)、皖豆20(S)、皖豆21(S)、皖豆22(S)、皖豆23(S)、皖豆26(S)、皖豆27(S)、皖豆28(S)、皖豆30(S)、皖豆31(S)、皖豆32(S)、文丰1号(S)、文丰2号(S)、文丰3号(S)、文丰4号(S)、文丰6号(S)、文丰7号(S)、文丰8号(S)、文丰9号(S)、徐豆1号(R)、徐豆2号(S)、徐豆3号(S)、徐豆4号(S)、徐豆7号(S)、徐豆8号(S)、徐豆9号(S)、徐豆10号(S)、徐豆11号(S)、徐豆12号(S)、徐豆13号(S)、徐豆14号(S)、徐豆15号(S)、徐豆16号(S)、徐豆17号(S)、徐豆18号(S)、徐豆126(S)、徐豆135(S)、跃进1号(S)、跃进2号(S)、跃进5号(S)、早黄2号(S)、早黄3号(S)、早熟1号(R)、早熟3号(R)、早熟6号、早熟7号(S)、中品12585(R)、中豆40(S)、丹豆1号(S)、丹豆3号(S)、丹豆4号(S)、东农51(S)、东农53(S)、汾豆79号(S)、汾豆95号、邯1049(S)、邯9013(S)、邯9133(S)、荷09-14(S)、合丰44(S)、合丰25(S)、济087201(R)、济J11124(S)、济J11130(S)、冀豆12(S)、晋遗34(S)、吉育94(R)、吉育97(S)、莒选23(S)、科丰1号(S)、科黄8号(S)、满仓金(S)、为民1号(S)、天隆1号(S)、铁丰3号(R)、烟豆4号(S)、烟黄3号(S)、早丰1号(S)、新大粒1号(R)、新黄豆(S)
含有WZInDel11目标片段的大豆品种
Soybean cultivar containing WZInDel11 target fragment		 豫豆23号(S)、郑92116(R)、周豆11(S)、皖豆24(S)、皖豆29(S)、皖宿2156(S)

表2

Williams和郑97196及其杂交衍生的F2:3群体对大豆疫霉分离物PsJS2抗性遗传分析"

品种和杂交组合
Parent and the cross		 世代
Generation		 数量
Sample size		 观察值Observed number 卡平方检验Chi squared tests
抗病
R		 杂合
Rs		 感病
S		 期望值
Expected ratio		 χ2 P
郑97196 Zheng 97196 P1 20 20 — —
Williams P2 20 — — 20
Williams×郑97196
Williams×Zheng 97196		 F2:3
 188
 47
 91
 50
 1:2:1
 0.29
 0.86

图1

RpsZheng定位区域遗传连锁图和物理图 A: 利用大豆3号染色体上与大豆抗疫霉病基因RpsZheng连锁的16个SSR标记构成的遗传连锁图。左侧是遗传距离(cM), 右侧是分子标记与RpsZheng的位置。B: RpsZheng定位区域的遗传连锁图谱。左侧是遗传距离(cM), 右侧是分子标记与RpsZheng的位置。C: InDel标记InDel7和InDel12之间构建的RpsZheng物理图谱。黑色部分表示在F2:3群体中没有发生重组交换的共分离区域。右侧为分子标记, 左侧为物理位置。"

表3

大豆3号染色体上RpsZheng定位区间内(105.2 kb)的基因及注释(Glycine max V2.0)"

基因模型名称
Gene model name		 物理位置
Position		 基因注释
Annotation
Glyma.03g035200 Gm03:4265953…4274479 锌指CW型卷曲螺旋结构域蛋白3 Zinc finger CW-type coiled-coil domain protein 3
Glyma.03g035300 Gm03:4274744…4278897 丝氨酸/苏氨酸蛋白激酶 Serine/threonine protein kinase
Glyma.03g035400 Gm03:4282985…4285957 五肽重复(PPR)超家族蛋白 Pentatricopeptide repeat (PPR) superfamily protein
Glyma.03g035500 Gm03:4289118…4294184 类氨基转移酶, 植物移动域家族蛋白
Aminotransferase-like, plant mobile domain family
Glyma.03g035600 Gm03:4295128…4296444 脂质转移蛋白1 Lipid transfer protein 1
Glyma.03g035700 Gm03:4309159…4309326 脂质转移蛋白5 Lipid transfer protein 5
Glyma.03g035800 Gm03:4317466…4321009 扩张素A2 Expansin A2
Glyma.03g035900 Gm03:4322884…4330292 富含MAC/Perforin结构域的蛋白质 MAC/Perforin domain-containing protein
Glyma.03g036000 Gm03:4335981…4341371 丝氨酸/苏氨酸蛋白激酶 Serine/threonine protein kinase
Glyma.03g036100 Gm03:4337336…4337912 无功能注释 No functional annotation
Glyma.03g036200 Gm03:4360966…4367363 多抗药性蛋白 Multidrug resistance protein

图2

标记WZInDel11在大豆品种中的分子检测结果 A: 标记WZInDel11对含有已知抗疫霉病基因的23个大豆品种检测, Williams和郑97196为对照; B: 标记WZInDel11在226个大豆品种中鉴定的含有目标片段的6个大豆品种的电泳图。"

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