Extraction and cloning of interleukine- 2 gene from the Iranian chicken

Subject Areas : Microbial Biotechnology

Hamideh Amini ¹ , Seyed Davoud Hoseini ² , Jamileh Nowroozi ³ , Delavar shahbazzadeh ⁴

1 - Department of Microbiology, Faculty of Biology, Islamic Azad University, North Tehran Branch, Tehran, Iran
2 - Department of Cellular and Molecular Biology, Razi Vaccination and Serum Research Institute, Arak, Iran
3 - Department of Microbiology, Faculty of Biology, Islamic Azad University, North Tehran Branch, Tehran, Iran
4 - Department of Biotechnology, Pasteur institute, Tehran, Iran

Received: 2013-04-14 Accepted : 2013-04-14 Published : 2010-04-01

Keywords: Adjuvant, Cloning, Chicken IL-2, recombinant vaccine, Cytokine,

Abstract :

Background and Objectives: Nowadays, the immune cytokines, such as the chicken IL-2 (chIL-2) gene are incorporated into vaccination regimens used by the poultry industry. The cytokines can potentially enhance the efficiency of vaccines according to increase T lymphocyte proliferation, B lymphocyte development and natural killer cell (NK) activation. The aim of this study was extraction and cloning of Iranian chicken interleukine-2 (chIL-2). Materials and Methods: First of all, in order to extract chIL-2 gene, total cell RNAs were isolated by culturing the harvested splenocytes and using Trizol reagent. The chIL-2 specific mRNA was converted into cDNA using reverse Transcriptase (RT) and specific designed primers. Then, The PCR product was ligated into the PTZ57R/T plasmid (TA-cloning kit) and was transformed into the competent Top10 E. coli using heat shock method. Results: A unique band of 668-bp was obtained after RT-PCR amplification. The results of restriction enzyme, colony PCR from transformed colonies and also gene sequencing confirmed the existence of desired gene in transformants bacteria. Conclusion: For the first time in Iran we could extract and clone the chIL-2 gene in bacteria and gene alignment. A direct sequencing test showed 99% similarity between this gene and the established data in NCBI.

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