持有Pi9基因的水稻单基因系IRBL9-W对稻瘟病菌苗期和成株期抗性差异  

Different Reactions of Rice Monogenic Line IRBL9-W Harboring Pi9 Gene to Magnaporthe oryzae Containing AvrPi9 During Seedling and Adult-plant Stages

作　　者：刘树芳[1,2] 董丽英 李迅东[1,2] 周伍民 杨勤忠[1,2] LIU Shufang;DONG Liying;LI Xundong;ZHOU Wumin;YANG Qinzhong(Agricultural Environment Resources Research Institute,Yunnan Academy of Agricultural Sciences/Key Laboratory of Green Prevention and Control of Agricultural Transboundary Pests of Yunnan Province,Kunming 650205,China;Scientific Observing and Experimental Station of Crop Pests in Kunming,Ministry of Agriculture and Rural Affairs,Kunming 650205,China;College of Agriculture,Yunnan University,Kunming 650091,China)

机构地区：[1]云南省农业科学院农业环境资源研究所/云南省农业跨境有害生物绿色防控重点实验室,昆明650205 [2]农业农村部昆明作物有害生物科学观测实验站,昆明650205 [3]云南大学农学院,昆明650091

出　　处：《中国水稻科学》2021年第3期303-310,共8页Chinese Journal of Rice Science

基　　金：云南省基础研究计划重点资助项目(202001AS070006);科技部国家重点研发专项(2016YFD0100101);国家自然科学基金资助项目(31860524);云南省农业科学院院基金资助项目(YJZ201803)。

摘　　要：【目的】Pi9是一个广谱稻瘟病抗性基因,田间病圃监测发现持有Pi9的水稻单基因系IRBL9-W在苗期高抗稻瘟病,但却感穗瘟。探明水稻单基因系IRBL9-W在苗期抗病而孕穗末期感染穗颈瘟的原因,为Pi9基因在水稻抗病育种中的有效利用提供参考。【方法】利用从IRBL9-W穗颈瘟病斑上分离的8个单孢菌株以及实验室保存的单孢菌株Y363,在温室分别对单基因系IRBL9-W苗期和孕穗末期进行接种鉴定;并利用病原菌AvrPi9基因的特异引物对9个单孢菌株进行PCR扩增及产物测序;提取水稻单基因系IRBL9-W苗期叶片和抽穗期穗部的总RNA,通过半定量RT-PCR以及实时qRT-PCR分析Pi9基因在苗期和穗期的表达。【结果】在温室人工喷雾接种条件下,IRBL9-W在苗期对从穗颈瘟上分离的8个单孢及对照菌株Y363均表现为抗病;随机选取的2个从IRBL9-W穗颈瘟病样分离的单孢菌株(YX2-7-1和YX2-15-1)及对照菌株Y363对孕穗末期IRBL9-W注射接种,接种的植株表现出典型的穗颈瘟症状;AvrPi9的等位基因分析结果表明,与AvrPi9相比,Y363中的等位基因与AvrPi9完全相同,而从IRBL9-W穗瘟分离的8个单孢菌株中编码区与AvrPi9基因完全相同,但在编码起始位置上游−264 bp处缺失16 bp的一段序列。由于IRBL9-W苗期对这些菌株均表现抗病,推测这段序列的缺失并不影响AvrPi9基因的功能;实时qRT-PCR分析结果表明,Pi9基因在穗部的表达量为苗期叶片表达量的47.3%。【结论】在水稻单基因系IRBL9-W中,与苗期叶片中Pi9基因的表达量相比,Pi9基因在穗部表达量的明显降低可能是造成IRBL9-W穗期感稻瘟病的原因。【Objective】The Pi9 gene confers broad-spectrum resistance against Magnaporthe oryzae.Rice monogenic line IRBL9-W carrying single blast resistance gene Pi9 is highly resistant to leaf blast during the seedling stage,but it is susceptible to neck blast at adult-plant stage in blast nursery.It is important to analyze the mechanism behind the reversion of resistance to Magnaporthe oryzae,which will provide important information for effective utilization of Pi9 in disease-resistant rice breeding program.【Method】Eight single spore strains of M.oryzae isolated from neck blast samples of diseased IRBL9-W,and one isolate Y363 were artificially inoculated by spraying at seedling stage to pathotype their pathogenicity on 24 monogenic lines including IRBL9-W.Two out of eight strains and control strain Y363 were further selected and inoculated on IRBL9-W at late-booting stage by injecting to evaluate their pathogenicity.The alleles of M.oryzae AvrPi9 gene cognate to rice blast resistance gene Pi9 were amplified and sequenced with AvrPi9-specific primers in nine strains.Total RNA of the rice monogenic line IRBL9-W in seedling and late-booting stage was extracted from young leaf and panicle,respectively.The expression analysis of Pi9 gene was performed by semi-quantitative RT-PCR and real-time qRT-PCR.【Result】The monogenic line IRBL9-W was completely resistant to the nine M.oryzae strains at the seedling stage exposed to the artificial spraying.However,IRBL9-W was susceptible to panicle blast when inoculated with both two randomly selected strains(YX2-7-1 and YX2-15-1)isolated from panicle blast lesions of IRBL9-W,and the control strain Y363 by injecting inoculation at the late-booting stage.Compared with AvrPi9,allelic sequence amplified from Y363 was identical to cloned AvrPi9,and the coding region of the other eight strains isolated from IRBL9-W panicle blast was identical to AvrPi9 gene,except for a 16-bp deletion at−264 bp upstream of the coding starting position in the eight strains.Given high resistance of

关 键 词：水稻 稻瘟病菌 Pi9 苗期 成株期 

分 类 号：S435.111.41[农业科学—农业昆虫与害虫防治]