Cloning of the Recombinant Cytochrome P450 Cyp141 Protein of Mycobacterium tuberculosis as a Diagnostic Target and Vaccine Candidate

Iranian Red Crescent Medical Journal
Mohammad Rabiee-FaradonbehAbolfazl Gholipour

Abstract

Tuberculosis has been announced as a global emergency by World Health Organization and the second infectious agent of mortality worldwide. The general policy in the development of new vaccines is to develop some vaccines with higher efficiency not only for infants but also for adults compared with the Bacillus Calmette-Guerin vaccine. Recently, cytochrome P450 cyp141 has been introduced as a new target for detecting Mycobacterium tuberculosis from clinical samples. The aim of this study was to clone this gene in order to pave the way for more evaluation. M. tuberculosis H37Rv DNA was extracted by a standard phenol-chlorophorm protocol. After designing the specific primers, P450 cyp141 gene was replicated by PCR. The purified PCR products were then subcloned into the pTZ57R/T plasmid vector. After extraction, enzyme digestion, and recombinant pTZ57R/T-cyp141 plasmid vector sequencing, the aforementioned products were cloned into a pET-26b plasmid vector. Then, the recombinant pET26b-cyp141 plasmid molecules were transformed to Escherichia coli strain BL21 (DE3) using the transformation method. Next, the recombinant pET26b-cyp141 plasmids were purified and evaluated by the enzyme digestion analysis. The cloning of P450 cyp141 gen...Continue Reading

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Methods Mentioned

BETA
nucleic acid amplification
PCR
electrophoresis

Software Mentioned

ProPred
Primer Designer
DNASIS
Basic Alignment Search

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