Development of transformation systems for kailaan (Brassica oleracea var. alboglabra) and kai choi (Brassica juncea cv. Leaf mustard) is presented in this thesis. The procedures include infection of hypocotyl explants of 14-days old seedlings with Agrobacterium strain MOG301 carrying a disarmed binary vector pKM-Cry1Ab. The binary vector carries a cDNA encoding the Bacillus thuringiensis class 1A(b) crystal protein in the T-DNA region, β-glucuronidase (gus) and neomycin phosphotransferase II gene (npt-II) were also assembled in the T-DNA region as reporter gene and selectable marker gene respectively. After co-cultivation for 3 days, the hypocoty1 explants were recovered on medium without antibiotics. Kanamycin was introduced after 1 week of recovery. B. juncea was successfully transfo...[ Read more ]

Development of transformation systems for kailaan (Brassica oleracea var. alboglabra) and kai choi (Brassica juncea cv. Leaf mustard) is presented in this thesis. The procedures include infection of hypocotyl explants of 14-days old seedlings with Agrobacterium strain MOG301 carrying a disarmed binary vector pKM-Cry1Ab. The binary vector carries a cDNA encoding the Bacillus thuringiensis class 1A(b) crystal protein in the T-DNA region, β-glucuronidase (gus) and neomycin phosphotransferase II gene (npt-II) were also assembled in the T-DNA region as reporter gene and selectable marker gene respectively. After co-cultivation for 3 days, the hypocoty1 explants were recovered on medium without antibiotics. Kanamycin was introduced after 1 week of recovery. B. juncea was successfully transformed at 14.7% to 43.3% and fertile seeds were collected. Agrobacterium infection was inhibited by glucosinolates in B. alboglabra cultures; exogenous thioglucosidase was introduced to the pre-culture medium to exhaust glucosinolates at the cutting ends before B. alboglabra hypocotyls were infected with Agrobacterium. The use of exogenous thioglucosidase successfully initiated transformation. Tissues from transgenic plants showed positive results in GUS histochemical assay. PCR was performed to confirm the integration of T-DNA into the plant genome.