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Identification of dominant isolate of Potato virus S in Iran and heterologous expression of its coat protein
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Nahid Masoudi   , Ahmad Rouhibakhsh * , Mohammad Hassan Asareh , Masoud Naderpour , Davoud Koolivand |
| Department of Plant protection, College of Agriculture, University of Guilan, Rasht, Iran. , a_rouhibakhsh1966@yahoo.com |
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Abstract: (4975 Views) |
| Gene expression in bacteria have already been used to study the protein of viruses and to produce specific antibodies. In this research, potato samples were collected from eight provinces of Iran and were tested for Potato virus S, as one of the most destructive viruses of potato fields, by DAS-ELISA and RT-PCR. Dominant isolate of PVS was selected according to coat protein (CP) gene sequences following phylogenetic analysis. Full length CP gene was amplified, cloned and sequenced, then was digested from pJET1.2/blunt vector, ligated into the expression vector pET-28a and the construct (pET-28a:PVS:CP) was transformed into Escherichia coli BL21 strain. Gene expression was optimized by induction with 0.5, 1 and 2 mM final concentrations of IPTG for 3, 4 and 6 h and was verified by SDS-PAGE and Western blotting. Based on the nucleotide analysis, the Iranian isolates of PVS were divided into three groups that one group with 22 members known as the dominant group. The expression of recombinant CP of about 34 kDa was proved by western blotting. Induction by 1 mM IPTG for 4 h proved to be the most efficient method of expression. This is the first report of the expression of PVS CP gene, which is important for the preparation of anti-PVS antibody. |
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| Keywords: Potato virus S, Gene expression, Recombinant protein, Phylogenetic analysis, PVS |
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Full-Text [PDF 728 kb]
(1658 Downloads)
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Type of Study: Research |
Subject:
Microrganisms and Viruses
Received: 2018/12/15 | Accepted: 2019/02/6 | Published: 2019/02/25
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