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Not AvailableOne of the most severe viral diseases of banana worldwide is caused by banana bunchy top virus (BBTV). It is a nanovirus transmitted by the aphids Pentalonia nigronervosa. Currently, there are no strategies available in order to completely protect bananas against the BBTV virus. The improvement of banana resistance to BBTV disease through conventional breeding has not been successful and therefore an alternative approach such as genetic engineering system requires producing resistance banana cultivar. In this study, gene isolation and construction of RNAi vector for bunchy top disease management in hill banana was determined. The target of the RNAi is the Rep protein encoded by the BBTV-DNA1. It can be concluded that the isolated rep gene displayed 100% sequence identity to the BBTV rep gene of Indian isolate deposited in the NCBI database. This result shows that BBTV rep gene is highly conserved among the different isolates of BBTV that infect the banana in India. Hence, an attempt was made to construct the RNAi vector for BBTV rep gene and developed method for evaluate the silencing mechanism through injection of Agrobacterium transient expression system in BBTV infected hill banana plants Virupakshi (AAB) using RNAi technology to silence and suppress the rep gene of BBTVICA