Agrobacterium-mediated transfer of chitinase gene in apple (Malus x domestica Borkh.) rootstock MM106

Abstract

Agrobacterium tumefaciens strain LBA4404 harbouring the transforming vector pCAMBAR chi11 containing chitinase gene was used in transformation studies using leaf as an explant in apple rootstock MM106. Putative shoots were selected on regeneration medium containing 5 mg/l hygromycin and 500 mg/l cefotaxime. Two-day preculturing and 96 h co-cultivation incubation were found effective to obtain maximum number of hygromycinresistant shoots. Two transgenic lines were confirmed after amplifying the integrated chi11 gene. The amplified fragments were further analysed for their authenticity using sequencing analysis. The obtained sequences were subjected to BLAST analysis, and showed 100 per cent homology with the rice endochitinase class I gene thus proves its integration. The transformation efficiency of putative transgenic shoots was observed to be 1.57 per cent.