分枝杆菌噬菌体CJAUS9 lysB基因的克隆与序列分析

Cloning and Sequence Analysis of Mycobacteriophage CJAUS9 lysB

关键词:  分枝杆菌  肌尾噬菌体  lysB  克隆  序列分析

Key words:  mycobacterium  myovirus  lysB  gene cloning  sequence analysis

摘要:

鉴于裂解酶可作为新型杀菌制剂被用于解决分枝杆菌的危害和耐药性问题，且尚未见有关于分枝杆菌噬菌体国内分离株裂解酶的研究。因此参照GenBank中lysB基因序列设计特异引物，以耻垢分枝杆菌噬菌体CJAUS9的基因组为模板，PCR扩增出1 029bp的条带，经酶切、PCR鉴定含有目的基因的克隆载体pMD19-T正确后进行测序分析。结果克隆出的lysB基因与已知分枝杆菌肌尾噬菌体lysB序列同源性为98.7%~99.1%；编码的氨基酸同源性为99%~100%，有酯酶保守的G-X-S-X-G基序，为后续研究LysB蛋白的生物学活性奠定了基础。

Abstract:

In view of development and application for lyase to solve questions of serious harmness and emergence of drug-resistant strains of mycobacteria disease. And there have not any research about lyase of mycobacteriophage isolated in domestic.Specific primers were designed according to sequences of myoviridae mycobacteriophage mycolylarabinogalactan esterase gene (lysB) in GenBank. Target bands were amplified by PCR with Mycobacterium smegmatis phage CJAUS9 genome as a template, connected with the cloning vector pMD19-T and transformed into E.coli DH5α. Recombinant plasmids were extracted, digested, identificated by PCR and sequenced. Results show 1 029bp lysB gene was successfully cloned and 98.7% to 99.1% sequence homology with other myoviridae mycobacteriophages’s lysB and 99% to 100% amino acid homology. GXSXG esterase conservative motif was also found. The research laid the foundation for study of biologic activity of LysB.