Designing of siRNAs for various target genes of Cucumber mosaic virussubgroup IB

Abstract

Cucumber mosaic virus (CMV) is a major production constraint in black pepper causing stunted disease. Resistant varieties are unavailable and control measures are not effective till now. Recently, RNA interference (RNAi) is the most promising strategy for combating virus infection in plants but the effectiveness depends on sequence specificity between the transgene and the targeting virus. This study was undertaken to design the most suitable region for double stranded RNA synthesis with maximum specificity and minimized off–targets for all the five genes of CMV from black pepper. A 400 bp off–target minimized region identified from each of the five genes was subjected to sequence polymorphism study with selected CMV subgroup IB strains and common ‘siRNAs’ were designed in silico. As 3b gene had the least variations (of 17%) with four common and potential siRNAs designed from this region in silico, a hairpin construct was assembled using this region in Agrobacterium that can be used for developing black pepper resistant to selected CMV subgroup IB strains.