TRANSFORMATION OF THE BANANA CULTIVAR 'NANJANGUD RASBALE' (SYN. 'RASTHALI', AAB, SILK SUBGROUP) WITH THE AMP GENE AND SCREENING FOR FUSARIUM RESISTANCE WITH A BIOASSAY

Abstract

The banana cultivar ‘Nanjangud Rasbale’ (syn. ‘Rasthali’, AAB, Silk subgroup) is highly susceptible to Fusarium wilt, caused by Fusarium oxysporum f. sp. cubense (Foc) race 1. This cultivar is facing extinction in its natural habitat. Transgenic plants of the cultivar were developed by transformation with the antimicrobial peptide (AMP) gene cloned from onion seeds and with constructs developed in pCAMBIA 2301. Apical meristems from in-vitro micropropagated plantlets were microwounded with naked gold particles, followed by co-cultivation with Agrobacterium containing the AMP gene. Rigorous selection in kanamycin 200 mg-1 took place for 8 months. Around 10% of the transformants that regenerated on the selection medium were further established. Integration of the genes npt II and AMP was confirmed by PCR, RT-PCR and Dot-blot technique. Plants regenerated from the suckers of these plants also tested positive for the presence of the gene by the above procedures. Regeneration and transformation percentage was 10% and 3%, respectively. The non-destructive, rapid bioassay developed by Companioni et al. (2003) was validated and used to screen for Fusarium resistance. Lesion development on leaves from sterile culture filtrates obtained from different incubation periods were studied, and an incubation period of 8 days was found to give maximum lesion development after inoculation. Leaves tested from 10-months old transformed plants had a lesion area of 1.61 mm2 ± 0.48 compared to controls with a lesion area of 6.24 mm2 ± 0.92. Transformants are being further multiplied for large-scale root challenging with Foc and further selection.