PCR Detection of Banana Bunchy Top Virus (BBTV) at Tissue Culture Level for the Production of Virus-free Planting Materials

Abstract

Banana bunchy top disease, caused by Banana bunchy top virus (BBTV), is a serious disease that affects the productivity of banana to a greater extent. The production of virus-free planting material is an efficient method in controlling this disease and increase productivity. In vitro propagation, especially meristem tip culture, has played a key role for obtaining a large number of virus free, homogenous planting materials in plantains and bananas (Musa spp.). In our previous study we have demonstrated that direct regeneration from male floral meristems is a rapid and simple method for clonal and mass propagation of Musa spp. cultivars ‘Virupakshi’ and ‘Sirumalai’ (AAB). Detection of BBTV by polymerase chain reaction (PCR) is very reliable and economically important for testing micropropagated banana plantlets for the absence of virus before release to farmers. In this study we have developed a protocol for early diagnosis of BBTV at tissue culture level itself through optimizing the Polymerase Chain Reaction (PCR) method. We designed the viral gene specific primers [coat protein (CP) gene and nuclear shuttle protein (NSP) gene] and standardize PCR conditions for detection of presence of BBTV. All the regenerated plantlets were tested for BBTV presence with standardized PCR and found to be completely free of virus. PCR tests are more sensitive than ELISA tests and are better to be suited to plant tissue with lower number of pathogens, such as in tissue culture conditions.