Regeneration and genetic transformation of hot pepper plants

The effective protocol for plant regeneration and genetic transformation, which can be used for the improvement of hot pepper (Capsicum annuum L.) quality, was established. High frequency of plant regeneration was observed when cotyledon segments were cultured on MS medium added with NAA (0.1 mg-L ^-1) and zeatin (2.0 mg-L^-1) or IBA (10.0 mg-L ^-1) and BAP (1.0 mg-L^-1). An addition of 5 μM AgNO ³ to these media improved the regeneration frequency up to 8%. Verification of presumed transgenic plants that survived in selection media was done by the amplification of selectable marker genes (ADA, NPTII, bar) by polymerase chain reaction (PCR). Successful expression of the ADA (adenosine deaminase) gene was detected by Northern blot and enzyme activity, suggesting a potential application of ADA gene as a selectable marker in genetic transformation system. Herbicide resistant gene, bar, was also introduced into hot pepper, and progenies of the transgenic plants were obtained. Stable inheritance (3:1 ratio) of bar gene was confirmed by PCR analysis and herbicide treatments of 5,000 mg-L'1. This highly efficient transformation system can be further used for the improvement of hot pepper quality using other useful genes.