Optimization of a biolistic transformation system for transfer of antifreeze gene KN2 and the bar herbicide resistance gene in common wheat.

We studied the effects of different media for callus induction and differentiation, and pre-culture period of immature wheat embryo culture on biolistic transformation efficiency for including antifreeze gene KN2 and bar conferring resistance to the herbicide bialaphos. The percentage of plantlets generated from induction and differentiation media without Cu2+ was lower than those cultured on differentiation media with Cu2+ (71.15%) or induction media with Cu2+ (68.45%) and both induction and differentiation media with Cu2+ (52.17%). The combinations of Nor medium for callus induction and Cu2+ medium for regeneration, and Cu2+ medium for induction and R medium for regeneration were superior for biolistic transformation. The calli induced on Cu2+ medium and pre-cultured for 4 d before biolistic transformation, and cultured on R medium after biolistic transformation produced the highest percentage (65%) of transgenic plantlets with the KN2 gene. Overall, about 50% plantlets regenerated from calli pre-cultured 4d before bombardment carried the KN2 gene; 44.7% of the plantlets carried the bar gene, which was higher than for any other treatment, followed by pre-culture 1d with 31.43% transformation rate for the KN2 gene and 20% transformation rate for the bar gene.