title: Cloning of the Recombinant Cytochrome P450 Cyp141 Protein of Mycobacterium tuberculosis as a Diagnostic Target and Vaccine Candidate.
abstract: BACKGROUND
  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.
  
  
  OBJECTIVE
  The aim of this study was to clone this gene in order to pave the way for more evaluation.
  
  
  METHODS
  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.
  
  
  RESULTS
  The cloning of P450 cyp141 gene was confirmed by the enzyme digestion and sequencing of the recombinant pTZ57R/T-cyp141 and pET26b-cyp141 plasmid vectors.
  
  
  CONCLUSIONS
  The results of this study demonstrated that the P450 cyp141 gene was successfully cloned into a pET26b plasmid vector as an expression vector. In this paper, for the first time in Iran, this gene was cloned for more purposes, including the expression and purification of the recombinant cytochrome P450 cyp141 protein.
authors.affiliation: Department of Microbiology and Immunology, Faculty of Medicine, Cellular and Molecular Research Center, Shahrekord University of Medical Sciences, Shahrekord, IR Iran ; Department of Microbiology, Shahrekord Branch, Islamic Azad University, Shahrekord,IR Iran.
authors.forename: Mohammad
authors.initials: M
authors.lastname: Rabiee-Faradonbeh
authors.affiliation: Department of Microbiology and Immunology, Faculty of Medicine, Alborz University of Medical Sciences, Karaj, IR Iran.
authors.forename: Davood
authors.initials: D
authors.lastname: Darban-Sarokhalil
authors.affiliation: Department of Microbiology, Faculty of Medicine, Tehran University of Medical Sciences, Tehran, IR Iran.
authors.forename: Mohammad Mehdi
authors.initials: MM
authors.lastname: Feizabadi
authors.affiliation: Department of Microbiology, Faculty of Medicine, Kermanshah University of Medical Sciences, Kermanshah, IR Iran.
authors.forename: Amirhooshang
authors.initials: A
authors.lastname: Alvandi
authors.affiliation: Department of Microbiology, Shahrekord Branch, Islamic Azad University, Shahrekord,IR Iran.
authors.forename: Hasan
authors.initials: H
authors.lastname: Momtaz
authors.affiliation: Department of Pathology, Al-Zahra Hospital, Isfahan University of Medical Sciences, Isfahan, IR Iran.
authors.forename: Neda
authors.initials: N
authors.lastname: Soleimani
authors.affiliation: Department of Microbiology and Immunology, Faculty of Medicine, Cellular and Molecular Research Center, Shahrekord University of Medical Sciences, Shahrekord, IR Iran.
authors.forename: Abolfazl
authors.initials: A
authors.lastname: Gholipour
keywords: Cloning; Cytochrome; Diagnostic; Mycobacterium; Tuberculosis