CLONING OF TOXOPLASMA GONDII GRANULAR ANTIGEN 5 (GRA5) IN EXPRESSION EUKARYOTIC PLASMID PCDNA3 AND ITS EXPRESSION ON CHO CELL

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NASERIFAR R.; GHAFFARIFAR F.; DALIMI ASL A.; SHARIFI Z.

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Journal: JOURNAL OF ILAM UNIVERSITY OF MEDICAL SCIENCES Year:2011 | Volume:19 | Issue:3 Page(s): 1-12

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Title

CLONING OF TOXOPLASMA GONDII GRANULAR ANTIGEN 5 (GRA5) IN EXPRESSION EUKARYOTIC PLASMID PCDNA3 AND ITS EXPRESSION ON CHO CELL

Author(s)

NASERIFAR R. | GHAFFARIFAR F. | DALIMI ASL A. | SHARIFI Z. | Issue Writer Certificate

Keywords

IMMUNIZATIONQ2

GRA5Q1

TOXOPLASMA GONDAIIQ2

N EXPRESSIO CELLQ2

DNA VACCINEQ2

Abstract

Introduction: Toxoplasmosis is a one of the most world-wide spread zoonosis representing a very serious clinical and veterinary problem. Toxoplasma gondii is an intracellular parasite that causes severe neurologic and ocular disease in immune compromised and congenitally infected individuals. The heavy incidence and severe or lethal damages of toxoplasmosis clearly indicate the need for the development of a more effective vaccine human vaccines are not available and current anti-toxoplasma treatment is disappointing. IMMUNIZATION with plasmid DNA, a relatively novel technique, is a promising vaccination technique. To improve the immune response by DNA vaccination various is one of the key for the success of the vaccine in the field. One of the most efficient ways to control this disease is IMMUNIZATION. However, so far, there is no effective vaccine available against this pathogen. An important source of human contamination with T. gondii is the consumption of raw or undercooked meat products. Toxoplasma gondii are widely prevalent in humans and other animals which can cause severe or lethal toxoplasmosis. So, the development of a more effective vaccine is needed urgently .Therefore, we prepare GRA5 plasmid to use as a vaccine.Materials & Methods: In this study, GRA5was cloned in pTZ57R; afterwards, it was transformed into TOP10 strain of E.coli Bacteria. The recombined plasmid extracted from E.coli bacteria and amplified through PCR technique. Besides, pcDNA3 Plasmid for receiving and cloning of GRA5 segment was digested by Hind3 & EcoRI enzymes. GRA5 was sub-cloned into pcDNA3 and the reaction ligation product was transformed for the above bacteria. The bacteria grew in LB culture with ampicillin. Recombined pcGRA5 plasmids were purified from E.coli by Plasmid extraction kit. Finally, recombinant plasmid using cell culture method was expressed in Cho cell.Findings: The accuracy of the results was confirmed by using restriction enzymes and PCR methods. GRA5 was cloned into expression eukaryotic plasmid pcDNA3. After sequencing pcGRA5 plasmid for cell expression Western blot method was used.Discussion & Conclusion: The results showed that cloning and transformation of fragment GRA5 in pcDNA3 was done properly.

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APA: Copy

NASERIFAR, R., GHAFFARIFAR, F., DALIMI ASL, A., & SHARIFI, Z.. (2011). CLONING OF TOXOPLASMA GONDII GRANULAR ANTIGEN 5 (GRA5) IN EXPRESSION EUKARYOTIC PLASMID PCDNA3 AND ITS EXPRESSION ON CHO CELL. JOURNAL OF ILAM UNIVERSITY OF MEDICAL SCIENCES, 19(3), 1-12. SID. https://sid.ir/paper/90225/en

Vancouver: Copy

NASERIFAR R., GHAFFARIFAR F., DALIMI ASL A., SHARIFI Z.. CLONING OF TOXOPLASMA GONDII GRANULAR ANTIGEN 5 (GRA5) IN EXPRESSION EUKARYOTIC PLASMID PCDNA3 AND ITS EXPRESSION ON CHO CELL. JOURNAL OF ILAM UNIVERSITY OF MEDICAL SCIENCES[Internet]. 2011;19(3):1-12. Available from: https://sid.ir/paper/90225/en

IEEE: Copy

R. NASERIFAR, F. GHAFFARIFAR, A. DALIMI ASL, and Z. SHARIFI, “CLONING OF TOXOPLASMA GONDII GRANULAR ANTIGEN 5 (GRA5) IN EXPRESSION EUKARYOTIC PLASMID PCDNA3 AND ITS EXPRESSION ON CHO CELL,” JOURNAL OF ILAM UNIVERSITY OF MEDICAL SCIENCES, vol. 19, no. 3, pp. 1–12, 2011, [Online]. Available: https://sid.ir/paper/90225/en

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