Agrobacterium mediated transformation of common bean (Phaseolus vulgaris L.)

Regeneration and shoot multiplication of common bean (Phaseolus vulgaris L. 'ICA Pijao') from half-cotyledonary nodes was achieved on modified Murashige and Skoog (1962) basal medium amended with 5 $ mu$M 6-benzylaminopurine. Histological studies confirmed the adventitious origin of the regenerated buds. Shoots were rooted ex vitro and developed into morphologically normal plants compared with seed-grown controls. The relative susceptibility of bean tissues to infection by a collection of wild-type Agrobacterium strains was tested. Positive transformation events were evaluated based on morphological and biochemical changes observed following Agrobacterium infection. The A. tumefaciens strain C58 was particularly virulent on greenhouse-grown plants, in vitro-derived stem sections, half-cotyledonary nodes and seedlings. A sensitive and rapid method was developed to detect opines using thin layer chromatography. Transient $ beta$-glucuronidase (GUS) gene expression was detected in 'ICA Pijao' bean buds regenerated from half-cotyledonary nodes following Agrobacterium-mediated gene transfer with the binary vector pGV1040 or p35SGUSINT. Four out of eight putative transformants contained the chimeric GUSINT gene following polymerase chain reaction (PCR) analysis. This was confirmed by Southern analysis of blotted PCR gels. However, there was no stable integration of the GUSINT gene as none of the R1 progeny showed an amplified GUSINT fragment with PCR.Doctor of Philosoph