EXPRESSION OF SUGARCANE STREAK MOSAIC VIRUS (SCSMV) COAT PROTEIN IN EXPRESSION VECTOR AS A FUSION PROTEIN WITH MALTOSE BINDING PROTEIN
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Authors
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R. Viswanathan
Sugarcane Breeding Institute
Coimbatore - 641007, Tamil Nadu, India
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R. Karuppaiah
Sugarcane Breeding Institute
Coimbatore - 641007, Tamil Nadu, India
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V. Ganesh Kumar
Sugarcane Breeding Institute
Coimbatore - 641007, Tamil Nadu, India
Abstract
The occurrence of Sugarcane streak mosaic virus (SCSMV), characterized as ‘Susmovirus’, a Potyviridae member, was recently reported in India. The virus as identified as one of the causative agents of mosaic in sugarcane along with Sugarcane mosaic virus (SCMV) either alone or in combination. Molecular diagnostic techniques were developed to detect both the viruses together in sugarcane and further studies were carried out to express the coat protein (CP) gene of SCMSV in E. coli system to produce recombinant antiserum. The complete SCSMVcp gene from mosaic infected sugarcane cv. Co 86032 was cloned using the primers SCSMV-F690 and SCSMV-R with BamHI and HindIII restriction sites. The viral protein was expressed as maltose binding protein (MBP)-SCSMVcp fusion protein in the expression vector pMAL c4x. The expressed fusion protein was purified using amylose resin column and eluted. Separation of purified fusion protein in 12% SDS-PAGE and subsequent Western blot with anti-MBP antiserum confirmed the authenticity of the expressed protein. Conventional virus purification is cumbersome, requires sophisticated instrumentation and does not eliminate other viruses and host protein contamination. The new recombinant expression of viral protein in in-vitro system has set a new platform to get large quantities of viral protein devoid of host contaminants. Further studies on serum production against the fusion protein and its validation are in progress.
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