Agrobacterium-mediated transformation of Brassica campestris ssp. parachinensis with synthetic Bacillus thuringiensis cry1Ab and cry1Ac genes.
An effective plant regeneration procedure and a gene transfer system via Agrobacterium tumefaciens were developed in Brassica campestris subsp. parachinensis [B. parachinensis]. Hypocotyls from 5-day-old seedlings with 2 days pre-culture were infected with Agrobacterium strain MOG301 harbouring a binary vector containing a synthetic Bacillus thuringiensiscry1Ab or cry1Ac gene with full codon-modification. After culture and selection on Murashige and Skoog medium supplemented with 4.0 mg/l BAP [benzyladenine], 2.0 mg/l NAA, 70 µsmallcap˜M AgNO3 and 50 mg/l kanamycin, a number of kanamycin-resistant plantlets were regenerated. Polymerase chain reaction and Southern blotting were used to identify and characterize the transgenic plants with the integrated cry1Ab or cry1Ac gene. Western blotting analysis of the transgenic plants confirmed the expression of insecticidal proteins encoded by cry1Ab or cry1Ac. Subsequent bioassay with larvae of the diamondback moth (Plutella xylostella) demonstrated that the transgenic plants were resistant to feeding damage.