Cloning of pe11 (LipX, Rv1169c) gene of Mycobacterium tuberculosis Beijing strain to pcDNA3.1 plasmid vector

IF 0.8 Q3 MULTIDISCIPLINARY SCIENCES
Lulut Azmi Supardi, Andriansjah Rukmana, Fithriyah Sjatha
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引用次数: 0

Abstract

Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis. It is a persistent global health problem with a high mortality rate. Currently, TB is controlled by administering the Bacillus Calmette-Guerin (BCG) vaccine, but the effectiveness of its protection varies among individuals in a population. The pe/ppe gene family comprises a typical group of genes that play a role in avoiding the host immune response and inducing persistent TB infection. Based on in silico analysis, the pe11 gene has estimated immunogenicity and potential as a TB seed vaccine candidate. The pe11 gene from an Indonesian isolate of an M. tuberculosis Beijing strain was amplified by polymerase chain reaction (PCR) and inserted into the mammalian expression vector pcDNA3.1. The recombinant vector pcDNA3.1-pe11 was used to transform Top10 competent Escherichia coli. Clones from the transformation were subjected to colony PCR to confirm the direction of the insert. Sequencing was performed to confirm the correctness of the insert sequence. In this study, the pe11 gene was successfully cloned into the pcDNA3.1 vector in the correct direction to assure PE11 expression. No mutations were found in the pe11 gene insert, compared with the M. tuberculosis H37Rv sequence as the standard. A pcDNA3.1 vector containing the pe11 gene derived from an M. tuberculosis Beijing strain was successfully constructed.
结核分枝杆菌北京株pe11 (LipX, Rv1169c)基因在pcDNA3.1质粒载体上的克隆
结核病(TB)是一种由结核分枝杆菌引起的传染病。这是一个长期存在的全球健康问题,死亡率很高。目前,结核病是通过接种卡介苗来控制的,但其保护效果因人群中的个体而异。pe/ppe基因家族包括一组典型的基因,在避免宿主免疫反应和诱导持续结核感染方面发挥作用。基于硅片分析,pe11基因估计具有免疫原性和作为结核种子疫苗候选物的潜力。采用聚合酶链反应(PCR)扩增印尼结核分枝杆菌北京株pe11基因,并将其插入哺乳动物表达载体pcDNA3.1中。利用重组载体pcDNA3.1-pe11转化Top10大肠杆菌。对转化后的克隆进行菌落PCR以确定插入物的方向。测序以确认插入序列的正确性。本研究成功将pe11基因沿正确方向克隆到pcDNA3.1载体中,保证了pe11的表达。与结核分枝杆菌H37Rv序列比较,pe11基因插入片段未发现突变。成功构建了结核分枝杆菌北京株pe11基因pcDNA3.1载体。
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来源期刊
Makara Journal of Science
Makara Journal of Science MULTIDISCIPLINARY SCIENCES-
CiteScore
1.30
自引率
20.00%
发文量
24
审稿时长
24 weeks
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