Regeneration and genetic transformation of chickpea

Chickpea genotypes ‘Pusa256’ and ‘PG186’ were tested for plant regeneration through multiple shoot production during 2002–04. The immature embryo and mature embryo axes were used as explants and cultured on medium containing MS macro salts, 4X MS micro salts, B5 vitamins, 3.0 mg/l BAP, 0.05 mg/l NAAand3%(w/v)sucrose. Optimumtransformationconditions were determined by treating explants with A. tumefaciens strain ‘LBA4404’ (pCAMBIA2301) containing the uid I and npt I genes. The cryl I insect resistance gene was cloned in pBinAR vector and the resulting vector was designated as pBinBt. The immature and mature embryo explants were transformed with A. tumefaciens strain LBA4404 (pBinBt). Kanamycin selection at 25 mg/l was initiated immediately after co-cultivation and the dose was doubled after 10 days. Multiple shoots were repeatedly selected on medium containing 50 mg/l kanamycin. The transgenic status of the kanamycin resistant shoots was confirmed by PCR analysis using gene specific primer sequences. Mature plants were recovered by grafting of the transformed shoots on to 2–3 week old seedlings of chickpea.