摘要
利用立枯丝核菌(Rhizoctonia solani)诱导胁迫下玉米高耐纹枯病自交系幼苗叶片所构建的cDNA文库中获得的EST序列,通过电子克隆和RT-PCR方法,结合cDNA末端快速扩增技术,从玉米叶片中分离克隆到衰老相关蛋白基因的全长cDNA序列,命名为ZmSAP(GenBank登录号:GU253311)。序列分析表明,ZmSAP全长1156bp,包含一个完整的603bp的开放阅读框(ORF),具有连续的Poly A尾和典型的加尾信号AATTAA。ProtParam程序预测表明该基因编码200个氨基酸,相对分子量为21.92kD,等电点为10.38。该氨基酸序列与豌豆、挪威云杉、寄生藻等有不同程度的同源性且在不同物种间具有一个保守结构域,染色体定位发现该基因位于玉米第1条染色体上。荧光定量PCR分析表明,在高耐纹枯病自交系R15和高感纹枯病材料478中ZmSAP基因在病菌胁迫初期呈诱导表达,可能参与了病原菌诱导的细胞程序性死亡过程,说明该基因与纹枯病菌胁迫响应机制密切相关。
Silico cloning,reverse transcription-polymerase chain reaction（RT-PCR）and rapid amplification of cDNA ends（RACE）were applied in cloning the senescence-associated protein gene with ESTs obtained from Rhizoctonia solani induced leaf cDNA library of R15 high resistant to banded leaf and sheath blight of maize as the basis.The full length cDNA of senescence-associated protein gene is 1 156 bp（GenBank accession mumber：GU253311）,which contains an open reading frame of 603 bp,a 5＇ untranslated region of 287 bp and a 3＇ untranslated region of 266 bp,and was named ZmSAP.ZmSAP has continuous poly A and a typical polyadenylation signal and putatively encodes a protein of 200 amino acid,with a predicted molecular weight of 21.92 kD and the isoelectric point of 10.38.The homology analysis indicated that the deduced amino acid sequence from zea mays L.shares different homology with those from Pisum sativum,Picea abies and Micromonas pusilla.Chromosome location revealed that this gene was located in the chromosome I.More-over,transcript analysis of ZmSAP gene to R.solani was examined in maize by real-time PCR.It was shown that the expression of ZmSAP was induced by R.solani in both the high resistant material R15 and the high susceptible line 478,and may be involved in the programmed cell death in response to R.solani.The experimental results indicated that ZmSAP is closely related to the response of the banded leaf and sheath blight in maize.
出处
《植物病理学报》
CAS
CSCD
北大核心
2010年第4期373-380,共8页
Acta Phytopathologica Sinica
基金
国家自然科学基金(30900901)
高产转基因玉米新品种培育(2008ZX08003-003)
四川省科技厅应用基础项目(2006J13-039)
四川省教育厅重点项目(07ZA063)
关键词
玉米
纹枯病
衰老相关蛋白
基因克隆
实时荧光定量PCR
maize
banded leaf and sheath blight（BLSB）
senescence-associated protein（SAP）
gene cloning
real-time PCR
