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:: Volume 23, Issue 7 (2-2016) ::
sjimu 2016, 23(7): 18-27 Back to browse issues page
Production of Recombinant Construct by Cloning of Protective Antigen Domain 4 Gene and Fusion of it with Lethal Factor Domain 1 Gene of Bacillus anthracis in E.coli
Mohammad Najarasl1, Mohammad Sadegh Hashemzadeh2, Hosein Honari 3, Jafar Mousavy1, Firouz Ebrahimi1, Hosein Pourhakkak1
1- Comprehensive University of Imam Hossein, Tehran, Iran.
2- Baqiyatallah University of Medical Sciences, Tehran, Iran.
3- Comprehensive University of Imam Hossein, Tehran, Iran. , msh.biotechnology@gmail.com
Abstract:   (5151 Views)

Introduction: Anthrax is a zoonotic disease. Bacterium Bacillus anthracis is the causative agent of the fatal disease. At present, the protective antigen (PA) is used as an effective vaccine against anthrax. Domain 4 of this antigen together with domain 1 of lethal factor (LF) are the potent immunogens of this bacteria and are as suitable candidates of vaccine against it. Our aim in this study is the cloning of protective antigen domain 4 (PAD4) genes and fusion of it with lethal factor domain 1 (LFD1) gene of the bacteria to evaluate their capability in protective immunity induction.

Materials & methods: In this experimental study, we used a recombinant pGEM-T easy vector containing LFD1 gene. Then PAD4 gene was amplified and isolated by PCR and cloned into another pGEM-T easy vector, separately. After that, ligation of PAD4 gene and LFD1 gene was done in mentioned vector with determination of LFD1 orientation and by PstI/XbaI restriction sites. The recombinant construct, resulted from these genes was sub-cloned into pET28a expression vector using BamHI/ XhoI restriction enzymes and after determination of genes orientation, the expression host BL21 was transformed by this recombinant vector.

Findings: First cloning and fusion of PAD4 and LFD1 gene fragments were successfully carried out in pGEM-T easy vector and after the confirmation of mentioned process by both of enzymatic digestion and PCR methods, the result recombinant construct was sub-cloned into pET28a.

Discussion & Conclusions: Since PAD4 and LFD1 are immunogenic regions, expression of the recombinant construct resulted from these ligated genes can be proposed as proper candidate of anthrax vaccine for induction of protective immunity.

Keywords: Bacillus anthracis, Protective antigen (PA), Lethal factor (LF), Fusion, Vaccine candidate
Full-Text [PDF 699 kb]   (1469 Downloads)    
Type of Study: Research | Subject: Bacteriology
Received: 2015/05/14 | Accepted: 2015/06/30 | Published: 2016/02/14
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Najarasl M, Hashemzadeh M S, Honari H, Mousavy J, Ebrahimi F, Pourhakkak H. Production of Recombinant Construct by Cloning of Protective Antigen Domain 4 Gene and Fusion of it with Lethal Factor Domain 1 Gene of Bacillus anthracis in E.coli. sjimu. 2016; 23 (7) :18-27
URL: http://sjimu.medilam.ac.ir/article-1-2702-en.html


Volume 23, Issue 7 (2-2016) Back to browse issues page
مجله علمی پزوهشی دانشگاه علوم پزشکی ایلام scientific journal of ilam university of medical sciences
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