Journal of Pure and Applied MicrobiologyVol. 9 No. 1

Detection of Sugarcane Mosaic Virus in Diseased Sugarcane using ELISA and RT-PCR Technique

A.S. Thorat1,3, R.K. Pal1, P.R. Shingote1,2*, S.B Kharte1,2, V.M. Nalavade1,2, D.R. Dhumale1, B.H. Pawar1 and K.H. Babu1

1Molecular Biology and Genetic Engineering Division, Vasantdada Sugar Institute, Manjari (Bk),Tal. Haveli, Pune - 412 307, India. 2Department of Biotechnology, Shivaji University, Kolhapur - 416004, India. 3Department of Botany, Shivaji University, Kolhapur - 416004, India.

Received on 06 November 2014 and accepted on 01 January 2014

 

ABSTRACT

Sugarcane mosaic virus (ScMV) causes yield losses in almost all the genotypes/clones of sugarcane. So the detection of ScMV is very important for disease management. In present study serological and molecular methods for rapid virus detections were used. Serological study including DAS-ELISA (Double Antibody Sandwich-Enzyme Linked Immunosorbent Assay) was optimized to identify the virus by using a monoclonal antibody. Leaf samples of sugarcane were collected from symptomatic and non-symptomatic plants. ELISA was done by sap extraction method. Total 80 cultivars of sugarcane were selected on the basis of morphological symptoms for ELISA test 70 % cultivars represented the presence of ScMV infection through ELISA. Two stepsRT-PCR (Reverse Transcriptase-Polymerase Chain Reaction) was performed for detecting ScMV using a pair of primers design to amplify a fragment in the coding region of ScMV coat protein. 93.75% cultivars showed the presence of ScMV infection through twostep RT-PCR. Further it was found that two step RT-PCR method is more stringent and will be used for the detection of low titer sample which were not detected by DAS-ELISA. In conclusion our findings suggest that ELISA and RT-PCR can be routinely used for ScMV detection with high efficiency of which RT-PCR is most reliable technique.

Keywords : Sugarcane genotypes, ScMV, DAS-ELISA, RT-PCR, serology.