稻瘟菌无毒基因AvrPii编码蛋白分子特性及转基因水稻的构建
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摘要
为了解稻瘟菌无毒基因AvrPii在水稻病原菌相关分子模式诱导的免疫反应(PTI)抗病分子信号调控中的功能,对AvrPii基因编码蛋白的分子结构特性进行分析,构建了AvrPii基因的植物表达载体幵进行遗传转化。结果表明:AvrPii蛋白由70个氨基酸组成,相对分子质量约为7 500,等电点为6.0,总平均亲水性值为–0.159;AvrPii蛋白包含1个信号肽、1个跨膜结构域和2个基序为LxAR和CxxCxxxxxxxxxxxH的保守结构域;利用农杆菌侵染转化法,获得AvrPii基因全长序列和不含信号肽AvrPiinsp的转基因株系13个,PCR扩增和实时荧光定量PCR鉴定结果表明,AvrPii和AvrPiinsp基因已成功转入受体品种日本晴幵得到稳定表达。
To characterize the biological function of AvrPii involved in rice PAMPs-triggered immunity(PTI) immune signaling,the properties of AvrPii protein encoded by AvrPii were analyzed,and the transgenic rice with AvrPii gene was constructed.The results showed that AvrPii protein is composed of 70 amino acids with a molecular weight of7 500,the theoretical isoelectric point is 6.0 and the average value of hydrophilic amino acids is –0.159.The AvrPii protein includes one signal peptide,one transmembrane domain,and two conserved motif with residues LxAR and Cxx CxxxxxxxxxxxH.Thirteen transgenic lines with full length AvrPii gene or AvrPiinsp gene without the signal sequence were obtained.PCR detection and expression analysis confirmed the insertion and the transcriptional expression of AvrPii and AvrPiinsp in the receptor cultivar Nipponbare.
To characterize the biological function of AvrPii involved in rice PAMPs-triggered immunity(PTI) immune signaling,the properties of AvrPii protein encoded by AvrPii were analyzed,and the transgenic rice with AvrPii gene was constructed.The results showed that AvrPii protein is composed of 70 amino acids with a molecular weight of7 500,the theoretical isoelectric point is 6.0 and the average value of hydrophilic amino acids is –0.159.The AvrPii protein includes one signal peptide,one transmembrane domain,and two conserved motif with residues LxAR and Cxx CxxxxxxxxxxxH.Thirteen transgenic lines with full length AvrPii gene or AvrPiinsp gene without the signal sequence were obtained.PCR detection and expression analysis confirmed the insertion and the transcriptional expression of AvrPii and AvrPiinsp in the receptor cultivar Nipponbare.
引文
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[2]LIU J,WANG X,MITCHELL T,et al.Recent progress and understanding of the molecular mechanisms of the rice-Magnaporthe oryzae interaction[J].Molecular Plant Pathology,2010,11:419-427.
[3]WANG B H,EBBOLE D,WANG Z H.The arms race between Magnaporthe oryzae and rice:diversity and interaction of Avr and R genes[J].J Integr Agric,2017,16:2746-2760.
[4]ZHAO H,WANG X,JIA Y,et al.The rice blast resistance gene Ptr encodes an atypical protein required for broad-spectrum disease resistance[J].Nat Commun,2018,9:2039.
[5]何峰,张浩,刘釐灵,等.水稻抗稻瘟病天然免疫机制及抗病育种新策略[J].遗传,2014,36(8):756-765.
[6]ORBACH M J,FARRALL L,SWEIGARD J A,et al.Atelomeric avirulence gene determines efficacy for the rice blast resistance gene Pi-ta[J].The Plant Cell,2000,12:2019-2032.
[7]KANZAKI H,YOSHIDA K,SAITOH H,et al.Arms race co-evolution of Magnaporthe oryzae AVR-Pik and rice Pik genes driven by their physical interactions[J].The Plant Journal,2012,72:894-907.
[8]CESARI S,THILLIEZ G,RIBOT C,et al.The rice resistance protein pair RGA4/RGA5 recognizes the Magnaporthe oryzae effectors AVR-Pia and AVR1-CO39by direct binding[J].The Plant Cell,2013,25:1463-1481.
[9]PARK C H,CHEN S,SHIRSEKAR G,et al.The Magnaporthe oryzae effector Avr Piz-t targets the RINGE3 ubiquitin ligase APIP6 to suppress pathogen-associated molecular pattern-triggered immunity in rice[J].The Plant Cell,2012,24:4748-4762.
[10]JONES J D,DANGL J L.The plant immune system[J].Nature,2006,444:323-329.
[11]YOSHIDA K,SAITOH H,FUJISAWA S,et al.Association genetics reveals three novel avirulence genes from the rice blast fungal pathogen Magnaporthe oryzae[J].The Plant Cell,2009,21:1573-1591.
[12]WAWRA S,TRUSCH F,MATENA A,et al.The Rx LRmotif of the host targeting effector AVR3a of phytophthora infestans is cleaved before secretion[J].The Plant Cell,2017,29:1184-1195.