Asian Journal of Microbiology, Biotechnology & Environmental Sciences Paper

Vol 8, Issue 3, 2006; Page No.(671-678)

PLANT REGENERATION AND AGROBACTER/L/M-MEDIATED GENE TRANSFER STUDIES IN STRAWBERRY TISSUES (FRAGARIA X ANANASSA DUCH.)

AMJAD M. HUSAINI AND DINESH K. SRIVASTAVA

Abstract

Plant regeneration and genetic transformation studies were conducted to standardize a protocol for high frequency plant regeneration and Agrobacierium-mediated gene transfer in strawberry tissues. Two types of explants viz. leaf and petiole were used. While using leaf as an explant, high frequency direct shoot regeneration was obtained on modified MS medium containing B5 vitamins, 2 % glucose and supplemented with 1.0 mg/I TDZ, 0.5 mg/1 IBA and 1.0 mg/I BA. High frequency direct shoot regeneration from petiole explant was obtained on modified MS medium containing B5 vitamins, 2% glucose and supplemented with 1.5 mg/1 BA and 0.5 mg/1 IBA. High percentage root regeneration in in vitro developed shoots was obtained on MS half strength medium when supplemented with 1.0 mg/I IBA. Regenerated plantlets of strawberry were transferred to the pots and acclimatized. Disarmed Agrobacterium tumefacicns LBA 4404 strain containing p-glucuronidase (gus) gene in binary vector pBI 121 along with a kanamycin resistance gene (npt-I1) for selection in both bacteria and plant was used for cocultivation experiment to transfer gus and tipt-11 genes in strawberry cells. After cocultivation, the tissues selected on kanamycin (50 mg/l) containing medium gave rise to transformed calli that were resistant to the antibiotic. Control explants were unable to grow on the selective medium. Transformation experiment could be scored as early as 4 weeks after selection. The presence of foreign gene was demonstrated in the transformed calli by expression of a chimeric bacterial gene that encodes B-glucuronidase.

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