小菜粉蝶颗粒体病毒颗粒体蛋白基因的序列测定及在大肠杆菌中的表达  

The Nucleotide Sequence and Expression in E. coli of the Pieris rapae Granulosis Virus Granulin Gene

作　　者：陈可[1] 张珈敏[1] 胡远扬[1] 刘传凤[1] 

机构地区：[1]武汉大学生命科学学院,湖北武汉430072

出　　处：《中国病毒学》2004年第5期493-497,共5页Virologica Sinica

摘　　要：根据颗粒体病毒颗粒体蛋白(Granulin)基因在其起始密码子上游的12个碱基高度保守序列(TATAAGGAATTT)以及大菜粉蝶颗粒体病毒(PbGV)的颗粒体蛋白基因的序列[1]设计引物,PCR扩增得到850bp左右大小的片段,核苷酸序列测定结果表明该病毒的granulin基因全长为855bp,起始密码位于第38～40位碱基,终止密码位于779～781位碱基,编码框序列全长为744;推测该基因编码一段由247个氨基酸组成的多肽,分子质量约为2.9178×104道尔顿.与其它颗粒体病毒颗粒体蛋白基因进行同源性比较,核苷酸同源性都在70%以上,氨基酸同源性都在75%以上,最高的为大菜粉蝶颗粒体病毒(PbGV),核苷酸同源性为97%,氨基酸同源性为98%.构建了重组表达载体pet-28a-Gran,IPTG诱导后经SDS-PAGE检测,表明获得了颗粒体蛋白基因在大肠杆菌BL21中的特异表达.According to the Granulin gene’s 12 highly conservative nucleotides (TATAAGGAATTT) upstream from the protein intiation codon and the sequence of the Pieris brassiae granulovirus (PbGV) granulin gene, primers were designed to amplify the Pieris rapae granulosis virus (PrGV) granulin gene. The product of PCR amplification was about 850bp. The result of sequencing the segment showed the complete nucleotide sequence of the segment was 855bp. The segment contained one ORF encoding PrGV granulin gene which starts at position 38～40 and ends at position 779～781. The length of the ORF is 744bp and the length of predicted protein is 247 amino acid with MW of about 2.9178×104D. Comparison of the identities with other granulosis virus granulin gene showed the granulin gene is highly conserved in granulosis virus. The PrGV granulin gene was inserted into an exprssion vector, PET-28a, to yield the recombinant expression plasmid pet-28a-Gran. After induction by IPTG, the result of SDS-PAGE showed the expression of PrGV granulin in E. coli was obtained.

关 键 词：小菜粉蝶颗粒体病毒 颗粒体蛋白 同源性比较 重组表达载体 

分 类 号：S476[农业科学—农业昆虫与害虫防治]