文献类型：期刊文章

作　　者：李文学[1] 李海峰[1] 金清洙[2]

机构地区：[1]延边大学农学院,吉林龙井133400 [2]吉林省延吉市动物卫生监督所,吉林延吉133400

出　　处：《Agricultural Science & Technology》

基　　金：Supported by Science and Technology Development Program of Jilin Province (20050703-4)~~

年　　份：2010

卷　　号：11

期　　号：5

起止页码：96-100

语　　种：中文

收录情况：CAB、CAS、CSA、CSA-PROQEUST、PROQUEST、普通刊

摘　　要：[Objective] The aim was to study cloning and prokaryotic expression of P23 major surface protein gene of Theileria sergenti. [Method] A pair of specific primers was designed according to the sequence of P23 major surface protein of T. sergenti (D84447).The P23 gene was amplified by PCR from genomic DNA of T. sergenti and cloned into pMD18-T vector to construct recombinant clonal vector pMD18-P23. Positive clones were identified by PCR screening and restriction digestion. A recombinant expression plasmid pGEX-4T-P23 was constructed by subcloning the cloned P23 gene into the linearized pGEX-4T-1 vector and transformed into E. coli BL21. After introduction by IPTG,the expressed fusion protein was identified by SDS-PAGE and Western-blotting. [Result] The cloned gene has a total length of 507 bp. Sequencing result showed that the nucleotide sequence of the cloned P23 gene shared 99.4% identity with that of P23 published in GenBank (D84447). The expressed fusion protein was 46 ku in molecular mass. Induction opportunity of zhours after culture inoculation was the best,the induction time of 6 h was the best,and induction temperature of 34 ℃ was the best as well,IPTG of 1 mmol/L had little effect on the expression. Western-blotting indicated that recombinant protein was recognized by specific antibody. [Conclusion] This study would lay a foundation for further research on the prevention and diagnose of T. sergenti.

关 键 词：Theileria sergenti P23 major surface protein gene Prokaryotic expression  

分 类 号：S858.23]