CLONING AND EVALUATION OF EXPRESSION OF A NOVEL OVERLAPPING REGION OF NS3 GENE OF HEPATITIS C VIRUS BY EXPRESSING VECTOR

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HOSSEINI SEYED YOUNES; SABAHI FARZANEH; MODARRESI MOHAMAD HOSSEIN; MOAZZENI SEYED MOHAMMAD; SABERI FIROOZI MEHDI; RAVANSHAD MEHRDAD; MEHRABANI DAVOOD; MOJARRAD MAJID

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Journal: Pathobiology Research Year:2011 | Volume:13 | Issue:4 Page(s): 21-32

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Title

CLONING AND EVALUATION OF EXPRESSION OF A NOVEL OVERLAPPING REGION OF NS3 GENE OF HEPATITIS C VIRUS BY EXPRESSING VECTOR

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Keywords

HCV VIRUSQ3

NS3 GENEQ3

DNA CONSTRUCTQ3

Abstract

Objective: In this project, our aim was to construct a novel expressing vector harboring a new sequence from overlapping region of NS3 GENE of HCV from infected Iranian patient.Materials and Methods: The partial NS3 (pNS3) gene was amplified by Nested-RT-PCR method using sera of HCV infected patients harboring genotype 1a. After purification and cloning the pNS3 into TA-cloning vector, the best colony was selected based on Blue/White screening and colony-PCR following by confirmation with sequencing and restriction digestion with BglII. The sequenced gene was compared with other reference sequences using alignment softwares. The resultant pNS3 gene subcloned into the expression vector, IRES vector, followed by selection the suitable clones by 2 different colony-PCRs. The gene expression was evaluated using GFP detection, RT-PCR and western blotting techniques after transfection of the IRES-pNS3 vector into the 293 cell line.Results: After pNS3 sequence amplification by RT-PCR, sequencing results showed high homology among the sequences with other reference sequences. This result also showed that it belonged to genotype 1 of HCV. Colony-PCR showed the insertion of gene into expressing vector with the right orientation. GFP expression, RT-PCR and western blotting confirmed transfection of vector, expression of pNS3 gene and production of its protein in 293 cells respectively.Conclusion: This novel expressing vector harboring partial region of NS3 GENE in compare to full NS3 GENE maybe more useful in immune induction by antigen presenting cells due to absence of genes responsible for protease activity of the protein in the setting of HCV vaccine.

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APA: Copy

HOSSEINI, SEYED YOUNES, SABAHI, FARZANEH, MODARRESI, MOHAMAD HOSSEIN, MOAZZENI, SEYED MOHAMMAD, SABERI FIROOZI, MEHDI, RAVANSHAD, MEHRDAD, MEHRABANI, DAVOOD, & MOJARRAD, MAJID. (2011). CLONING AND EVALUATION OF EXPRESSION OF A NOVEL OVERLAPPING REGION OF NS3 GENE OF HEPATITIS C VIRUS BY EXPRESSING VECTOR. PATHOBIOLOGY RESEARCH (MODARES JOURNAL OF MEDICAL SCIENCES), 13(4), 21-32. SID. https://sid.ir/paper/81024/en

Vancouver: Copy

HOSSEINI SEYED YOUNES, SABAHI FARZANEH, MODARRESI MOHAMAD HOSSEIN, MOAZZENI SEYED MOHAMMAD, SABERI FIROOZI MEHDI, RAVANSHAD MEHRDAD, MEHRABANI DAVOOD, MOJARRAD MAJID. CLONING AND EVALUATION OF EXPRESSION OF A NOVEL OVERLAPPING REGION OF NS3 GENE OF HEPATITIS C VIRUS BY EXPRESSING VECTOR. PATHOBIOLOGY RESEARCH (MODARES JOURNAL OF MEDICAL SCIENCES)[Internet]. 2011;13(4):21-32. Available from: https://sid.ir/paper/81024/en

IEEE: Copy

SEYED YOUNES HOSSEINI, FARZANEH SABAHI, MOHAMAD HOSSEIN MODARRESI, SEYED MOHAMMAD MOAZZENI, MEHDI SABERI FIROOZI, MEHRDAD RAVANSHAD, DAVOOD MEHRABANI, and MAJID MOJARRAD, “CLONING AND EVALUATION OF EXPRESSION OF A NOVEL OVERLAPPING REGION OF NS3 GENE OF HEPATITIS C VIRUS BY EXPRESSING VECTOR,” PATHOBIOLOGY RESEARCH (MODARES JOURNAL OF MEDICAL SCIENCES), vol. 13, no. 4, pp. 21–32, 2011, [Online]. Available: https://sid.ir/paper/81024/en

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