Characterization of Bt RNAi cotton plants produced through Agrobacterium-mediated genetic transformation for whitefly resistance
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Date
2024
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Punjab Agricultural University
Abstract
Cotton is India's important cash crop, and it plays a significant part in the country's industrial and agricultural economy. Whitefly (Bemisia tabaci) has been one of the most threatening polyphagous pests of cotton and no whitefly resistance germplasm is available in India. Also, there are no Bacillus thuringiensis (Bt) toxins known to be effective against whitefly. Generating RNAi cotton plants expressing dsRNA of vital gene of whitefly can be a novel approach to produce whitefly resistance in available Bt cotton variety. In present work, screening was done to check the presence of integrated RNAi cassette in genomic DNA of BtRNAi cotton plants in T1 generation. For this DNA from true leaves of BtRNAi cotton plants were isolated and PCR with RNAi cassette specific primers resulted in total seven positive plants. Non-chimeric nature of these seven BtRNAi cotton plants also confirmed through DNA-PCR in different branches which resulted in same 208 bp amplicon. Further expression of RNAi cassette (sense fragment of Vitellogenin gene) in seven PCR confirmed BtRNAi cotton plants (T1 generation) was seen. For this total RNA was isolated and cDNA was prepared, and expression of RNAi cassette (208 bp) was observed using qPCR. Upon insect bioassay fecundity rate on leaves of wild cotton plants was much higher (doubled) as compared to confirmed BtRNAi cotton plants and showed the effect of VG gene silencing in whiteflies which were fed on these plants. This proved the development of some tolerance/resistance against whitefly in these produced BtRNAi cotton plants through RNAi.
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Choudhary, Ankit Kumar (2024). Characterization of Bt RNAi cotton plants produced through Agrobacterium-mediated genetic transformation for whitefly resistance (Unpublished M.Sc. thesis). Punjab Agricultural University, Ludhiana, Punjab, India.