结核分枝杆菌H37Ra株CFP10基因的克隆与序列分析
Cloning and sequence analysis of the culture filter protein 10 gene from Mycobacterium tuberculosis H37Ra
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摘要: 目的:克隆结核分枝杆菌减毒株H37Ra培养滤液蛋白10(CFP10)基因,并对其进行序列分析。方法:自结核菌H37Ra株抽提基因组DNA为模板,以已知结核菌H37Ra株CFP10基因设计引物。通过聚合酶链反应(PCR)技术扩增出结核分枝杆菌H37Ra株CFP10基因,将其克隆至pTG19-T载体后,经PCR和双酶切鉴定后进行DNA序列测定,并以BLAST在线软件进行序列比对分析。结果:PCR产物电泳显示扩增片段约为300 bp,测序获得的片段开放编码框由303 bp组成,与H37Rv株CFP10基因同源性为100%,推导出编码氨基酸序列同源性为100%。结论:成功克隆结核分枝杆菌H37Ra株CFP10基因,其基因序列与H37Rv株CFP10基因序列无差异。Abstract: Objective: To obtain the culture filter protein 10( CFP10) gene from Mycobacterium tuberculosis H37Ra and clone it into plasmid for nucleotide sequence analysis. Methods: The genomic DNA from Mycobacterium tuberculosis H37Ra was extracted as template,and the primer was designed according to the CFP10 gene of Mycobacterium tuberculosis H37Ra. The CFP10 gene was amplified from the genome of Mycobacterium tuberculosis H37Ra by polymerase chain reaction( PCR) and cloned into pTG19-T vector. After identification by PCR and restriction enzyme analysis,the CFP10 gene was sequenced,analyzed and compared with that of H37Ra reported in GenBank. Results: The sequencing result showed that the gene was obtained with an open reading frame of 303 bp. Compared with the DNA sequence of Mycobacterium tuberculosis H37Ra,the homology was 100%,which deduced that the amino acid sequence of CFP10 was 100% identity. Conclusions: The CFP10 gene of Mycobacterium tuberculosis H37Ra had been cloned successfully and the homology was 100% compared with Mycobacterium tuberculosis H37Ra.
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