家蚕核型多角体病毒(陕西株)orf126基因的表达及与几丁质的体外结合实验

Expression of orf126 Gene of Bombyx mori Nucleopolyhedrovirus (Shaanxi Isolate) and in vitro Binding Assay of the Expressed Product with Chitin

为研究家蚕核型多角体病毒(BmNPV)orf126基因(Bm126)在感染宿主中的作用机制,采用PCR方法从BmNPV陕西株中克隆了切除N端23个氨基酸(预测信号肽序列)的Bm126基因,并将其和谷胱甘肽巯基转移酶(GST)融合后在大肠杆菌中进行了表达。利用纯化的BM126重组蛋白与几丁质进行体外结合实验,Western blot分析没有检测到BM126重组蛋白与几丁质结合的复合物,推测原核表达的重组BM126融合蛋白在体外不能与几丁质结合。

In order to study the functional mechanism of orf126 gene （Bm126） of Bombyx mori nucleopolyhedrovirus （BmNPV） during infecting the host, the Bm126 gene excising the N-terminal 23 amino acids of predicted signal peptide sequence was amplified by PCR from BmNPV Shaanxi isolate and was fused with glutathione S-transferase （GST） gene for expression in E. coli. in vitro binding assay was conducted for the purified recombinant BM126 protein with chitin. Western blot analysis did not identify any complex formed between the recombinant BM126 protein and chitin, suggesting that the prokaryotic expressed recombinant BM126 fusion protein is unable to bind chitin in vitro.