长江下游粳稻稻瘟病广谱抗性基因组合模式分析

doi:10.3864/j.issn.0578-1752.2021.09.006

中国农业科学 ›› 2021, Vol. 54 ›› Issue (9): 1881-1893.doi: 10.3864/j.issn.0578-1752.2021.09.006

长江下游粳稻稻瘟病广谱抗性基因组合模式分析

吴云雨^1,²(),肖宁^1,^2,⁴,余玲^1,²,蔡跃^1,²,潘存红^1,³,李育红^1,²,张小祥^1,²,黄年生^1,²,季红娟^1,²,戴正元^1,³,李爱宏^1,^2,³()

¹江苏里下河地区农业科学研究所,江苏扬州225007
²扬州大学江苏省作物基因组学和分子育种重点实验室,江苏扬州225009
³扬州大学江苏省粮食作物现代产业技术协同创新中心,江苏扬州225009
⁴中国农业科学院植物保护研究所植物病虫害生物学国家重点实验室,北京 100193

收稿日期:2020-07-13 接受日期:2020-08-24 出版日期:2021-05-01 发布日期:2021-05-10
通讯作者: 李爱宏
作者简介:吴云雨,E-mail：wuyunyuyu@163.com。
基金资助:
国家自然科学基金(31801342);国家自然科学基金(31971868);江苏省农业科技自主创新基金(CX182022);江苏省重点研发计划(BE2019339);江苏省农业重大新品种创制项目(PZCZ201702);江苏省作物基因组学和分子育种重点实验室(BM2018003);江苏省自然科学基金(BK20181216);中国农业科学院植物病虫害生物学国家重点实验室开放基金(SKLOF201909)

Construction and Analysis of Broad-Spectrum Resistance Gene Combination Pattern for Japonica Rice in Lower Region of the Yangtze River, China

WU YunYu^1,²(),XIAO Ning^1,^2,⁴,YU Ling^1,²,CAI Yue^1,²,PAN CunHong^1,³,LI YuHong^1,²,ZHANG XiaoXiang^1,²,HUANG NianSheng^1,²,JI HongJuan^1,²,DAI ZhengYuan^1,³,LI AiHong^1,^2,³()

¹Lixiahe Institute of Agricultural Sciences of Jiangsu, Yangzhou 225007, Jiangsu
²Jiangsu Key Laboratory of Crop Genomics and Molecular Breeding, Yangzhou University, Yangzhou 225009, Jiangsu
³Jiangsu Co-Innovation Center for Modern Production Technology of Grain Crops, Yangzhou University, Yangzhou 225009, Jiangsu
⁴State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193

Received:2020-07-13 Accepted:2020-08-24 Online:2021-05-01 Published:2021-05-10
Contact: AiHong LI

摘要/Abstract

摘要：

【目的】基因聚合是实现水稻稻瘟病广谱抗性的有效途径之一。通过构建粳稻背景下不同双基因聚合系,利用长江下游粳型稻瘟病菌（Magnaporthe oryzea）菌株评价其抗性效应并解析其抗性效应产生的构成因子,为长江下游粳稻抗稻瘟病育种提供广谱抗性基因组合模式和种质资源。【方法】以粳稻07GY31为背景的Piz基因座不同复等位基因（Pigm、Pi40、Pi9、Pi2、Pizt和Piz）单基因系为核心,利用不完全NCII交配设计,分别与其他广谱抗性基因（Pi1、Pi54和Pi33）单基因系杂交,经分子标记辅助选择和农艺性状筛选,共构建18种不同基因组合的双基因聚合系。2019年利用长江下游粳稻种植区采集、分离的109个稻瘟病代表性菌株进行苗瘟、穗瘟人工接种鉴定及不同病圃的自然诱发鉴定,评价不同双基因聚合系的抗性效应,并分析双基因聚合系抗性效应的构成因子。【结果】Genotyping by sequencing（GBS）分析表明所构建的双基因聚合系均具有较高的背景恢复率,分布于97.08%（PPL^Piz/Pi33）—99.08%（PPL^Pigm/Pi1）。表明除了目标基因区域不同外,所有双基因聚合系的遗传背景几乎完全与受体亲本07GY31一致。同时人工接菌鉴定表明绝大部分双基因聚合系苗瘟和穗瘟抗性水平都优于单基因系。其中苗瘟抗性效应较好的聚合系分别为PPL^Pigm/Pi1、PPL^Pigm/Pi54、PPL^Pigm/Pi33、PPL^Pi9/Pi33、PPL^Pi9/Pi54、PPL^Pi40/Pi54、PPL^Pi40/Pi33、PPL^Pi40/Pi1、PPL^Pi9/Pi1, 而穗瘟抗性效应较好的聚合系分别为PPL^Pigm/Pi1、PPL^Pigm/Pi54、PPL^Pigm/Pi33、PPL^Pi40/Pi33、PPL^Pi40/Pi54、PPL^Pi40/Pi1、PPL^Pizt/Pi33。不同抗性基因聚合后产生不同的效应,其中互补效应高且能有效表达是提高双基因聚合系苗瘟和穗瘟抗性的关键因子。双基因聚合系PPL^{Pigm / Pi1}、PPL^{Pigm / Pi54}和PPL^{Pigm / Pi33}在苗瘟和穗瘟的人工接种,以及在不同病圃的自然诱发鉴定中均表现稳定的广谱抗性,同时,农艺性状调查结果也表明这3个双基因聚合系的基本农艺性状与轮回亲本07GY31基本一致,因此,基因组合Pigm/Pi1、Pigm/Pi54和Pigm/Pi33是适于长江下游粳稻的广谱抗性基因组合模式。【结论】抗性基因的组合方式影响聚合系的抗性水平,互补效应高且能有效表达是粳型双基因聚合系抗性效应提高的关键因子。本研究构建的双基因聚合系及其抗性效应分析为长江下游广谱稻瘟病抗性粳稻品种的精准培育提供了种质资源和理论支撑。

关键词: 长江下游, 粳稻, 稻瘟病菌, 稻瘟病, 基因聚合, 效应分析

Abstract:

【Objective】Gene pyramiding is one of the most effective ways to achieve broad-spectrum resistance against Magnaporthe oryzae. The objective of this study is to construct a set of polygene pyramiding lines (PPLs) under the background of japonica rice, to evaluate their resistance performances and analysis the components of their resistance effects using M. oryzae strains collected from lower region of the Yangtze River, China, thus providing broad-spectrum resistance gene combination pattern and germplasm resources for japonica rice resistance breeding in lower region of the Yangtze River, China. 【Method】Monogenic lines with multiple alleles of the Piz locus (Pigm, Pi40, Pi9, Pi2, Pizt and Piz) with the background of japonica rice 07GY31 as the backbone, crossed with other broad-spectrum resistance gene Pi1, Pi54 and Pi33, respectively using the incomplete NCII mating design. A total of 18 different PPLs were constructed using marker-assisted selection (MAS) and agronomic traits screening. Artificial inoculation assays at seedling and heading stage with 109 representative M. oryzae strains collected from lower region of the Yangtze River, combined with natural induction identification under multiple field environments were conducted to evaluate the resistance performances of different PPLs, and to analyze the component factors of the resistance effects of the PPLs. 【Result】Genotyping by sequencing (GBS) analysis shows that the constructed PPLs all have a high background recovery rate, which was ranging from 97.08% (PPL ^Piz/Pi33) to 99.08% (PPL^Pigm/Pi1), indicated that the genetic background of all PPLs was almost fully identical to that of the recurrent parent 07GY31. The seedling blast and panicle blast resistance levels of most PPLs were significantly higher than those of monogenic lines under artificial inoculation conditions, the PPLs with better resistance to seedling blast are PPL^Pigm/Pi1, PPL^Pigm/Pi54, PPL^Pigm/Pi33, PPL^Pi9/Pi33, PPL^Pi9/Pi54, PPL^Pi40/Pi54, PPL^Pi40/Pi33, PPL^Pi40/Pi1 and PPL^Pi9/Pi1, respectively, and the PPLs with outstanding performance in panicle blast are PPL^Pigm/Pi1, PPL^Pigm/Pi54, PPL^Pigm/Pi33, PPL^Pi40/Pi33, PL^Pi40/Pi54, PPL^Pi40/Pi1 and PPL^Pizt/Pi33, respectively. Different resistance gene combinations produced different effects after pyramiding. High complementary effect and which could be fully expressed is the key factor for the improvement of the resistance level of seedling blast and panicle blast of the PPLs. In addition, PPL^Pigm/Pi1, PPL^Pigm/Pi54 and PPL^Pigm/Pi33 displayed broad-spectrum resistance in artificial inoculation at seedling and heading stage, and showed stable broad-spectrum resistance under different disease nurseries. Besides, agronomic traits evaluation also showed PPLs with these three gene combinations were at par to the recurrent parent. Therefore, Pigm/Pi1, Pigm/Pi54 and Pigm/Pi33 are broad-spectrum resistance gene combination patterns suitable for japonica rice resistance breeding in lower region of the Yangtze River, China. 【Conclusion】The combination pattern of resistance genes affects the resistance level of the PPLs, and high complementary effect and which could be fully expressed is the key factor for the improvement of the resistance level of the PPLs in japonica background. In addition, the development of PPLs and component factors analysis in this study provides valuable theoretical support and innovative germplasm resources for the precise breeding broad-spectrum japonica varieties in lower region of the Yangtze River, China.

Key words: lower region of the Yangtze River, japonica rice, Magnaporthe oryzae, rice blast, gene pyramiding, effect analysis

吴云雨,肖宁,余玲,蔡跃,潘存红,李育红,张小祥,黄年生,季红娟,戴正元,李爱宏. 长江下游粳稻稻瘟病广谱抗性基因组合模式分析[J]. 中国农业科学, 2021, 54(9): 1881-1893.

WU YunYu,XIAO Ning,YU Ling,CAI Yue,PAN CunHong,LI YuHong,ZHANG XiaoXiang,HUANG NianSheng,JI HongJuan,DAI ZhengYuan,LI AiHong. Construction and Analysis of Broad-Spectrum Resistance Gene Combination Pattern for Japonica Rice in Lower Region of the Yangtze River, China[J]. Scientia Agricultura Sinica, 2021, 54(9): 1881-1893.

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https://www.chinaagrisci.com/CN/Y2021/V54/I9/1881

图/表 6

表1

图1

图2

图3

表2

18个双基因聚合系及轮回亲本07GY31的基本农艺性状表现"

品系 Line	抽穗期 Heading date (d)	株高 Plant height (cm)	单株有效穗数 Number of panicles per plant	每穗总粒数 Number of grains per panicle	结实率 Seed fertility (%)	千粒重 1000-grain weight (g)	单株产量 Grain yield per plant
PPL^Pigm/Pi1	97.2±1.26a	100.55±3.05a	11.38±1.32a	126.32±4.58a	92.02±0.35a	26.32±1.42a	34.81±2.02a
PPL^Pigm/Pi54	96.5±0.24a	99.82±2.58a	11.22±1.04a	126.25±6.72a	92.52±0.82a	26.04±1.08a	34.12±1.56a
PPL^Pigm/Pi33	97.0±2.13a	99.65±2.85a	10.78±0.95a	127.95±5.56a	91.89±0.64a	26.54±0.84a	33.63±1.88a
PPL^Pi40/Pi1	95.5±2.28a	98.75±0.85a	10.95±1.58a	125.85±7.22a	93.25±1.08a	27.06±1.32a	34.77±3.14a
PPL^Pi40/Pi54	94.7±1.89a	98.28±1.42a	11.32±0.68a	127.46±8.18a	94.28±0.92a	26.38±1.62a	35.89±2.28a
PPL^Pi40/Pi33	95.2±1.56a	99.12±1.78a	10.56±1.84a	126.28±5.42a	93.75±0.48a	27.59±0.82a	34.49±1.92a
PPL^Pi9/Pi1	91.8±2.32b	98.30±1.65a	11.24±1.08a	120.24±3.92b	88.32±0.74b	25.88±0.82a	30.89±0.89ab
PPL^Pi9/Pi54	90.4±1.74b	99.18±1.58a	10.68±0.82a	118.65±7.25b	87.45±0.98b	26.12±1.28a	28.94±1.02b
PPL^Pi9/Pi33	91.5±1.82b	98.86±0.92a	11.36±1.38a	119.84±6.26b	88.26±1.05b	26.16±0.85a	31.43±1.65ab
PPL^Pi2/Pi1	98.4±2.16a	102.45±2.37ab	11.69±1.32a	125.38±4.38a	92.45±0.85a	26.64±1.64a	36.10±1.32a
PPL^Pi2/Pi54	97.5±1.84a	101.84±2.72a	11.88±1.15a	126.56±9.82a	93.28±0.62a	25.98±1.25a	36.43±1.74a
PPL^Pi2/Pi33	97.2±1.45a	103.22±1.84b	11.21±0.75a	125.58±10.52a	93.75±1.12a	26.49±0.92a	34.96±2.84a
PPL^Pizt/Pi1	100.5±2.23c	99.48±2.24a	10.95±1.21a	128.25±6.69a	93.55±1.43a	27.18±1.36a	35.71±2.25a
PPL^Pizt/Pi54	101.8±0.95c	98.95±3.21a	11.22±1.43a	127.84±8.82a	92.45±0.98a	26.52±1.52a	35.16±1.95a
PPL^Pizt/Pi33	99.6±1.48a	100.24±2.88a	11.47±1.28a	126.57±4.78a	93.12±1.28a	26.35±0.79a	35.62±1.35a
PPL^Piz/Pi1	97.5±2.32a	97.32±1.69a	10.98±0.92a	128.48±5.92a	92.88±0.85a	26.38±0.88a	34.56±0.85a
PPL^Piz/Pi54	96.5±2.65a	98.48±1.86a	11.22±0.78a	126.32±6.74a	92.05±0.58a	25.94±1.35a	33.84±1.63a
PPL^Piz/Pi33	97.2±2.18a	97.85±0.82a	10.92±1.26a	127.62±7.28a	92.69±0.82a	26.36±1.02a	34.05±1.28a
07GY31	96.8±1.24a	98.55±2.52a	11.24±1.34a	126.57±6.28a	92.36±0.74a	26.28±1.28a	34.53±1.56a

表2

图4

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基因 Gene	标记 Marker	引物序列 Primer sequence (5′-3′)	退火温度 Annealing temperature (℃)	预期片段大小 Expected size (bp)
Pi2	Pi2-InDel	F: GCAGCGGCTAGGGTTTATC	60	110
		R: CACCCAGCAACTGATTTGTCA
Pi9	M-Pi9	F: GCTGTGCTCCAAATGAGGAT	55	291
		R: GCGATCTCACATCCTTTGCT
Pi40	AP5659-5	F: CTCCTTCAGCTGCTCCTC	55	288
		R: TGATGACTTCCAAACGGTAG
Pigm	Z4794	F: TGAATGTGAGAGGTTGACTGTGG	55	334
		R: CACGCCACCCTTCAATGGAGACT
Pizt	AP22	F: GTGCATGAGTCCAGCTCAAA	58	143
		R: GTGTACTCCCATGGCTGCTC
Piz	S29742	F: CAGTGAAACGAACGCTATG	55	454
		R: AATAGGAAGGGTTGATGTTG
Pi1	RM224	F: ATCGATCGATCTTCACGAGG	55	163
		R: TGCTATAAAAGGCATTCGGG
Pi54	Pi54-1	F: ATCGATCGATCTTCACGAGG	55	216
		R: GCTTCAATCACTGCTAGACC
Pi33	RM72	F: CCGGCGATAAAACAATGAG	55	240
		R: GCATCGGTCCTAACTAAGGG

长江下游粳稻稻瘟病广谱抗性基因组合模式分析

Construction and Analysis of Broad-Spectrum Resistance Gene Combination Pattern for Japonica Rice in Lower Region of the Yangtze River, China

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