Agrobacterium-mediated gene transfer in papaya
1991
Petcharat Siriwong
Chimeric genes consisting of CaMV 35S promoter/GUS/NOS/NPT-II in the binary vector were transferred into papaya tissues via Agrobacterium tumefaciens with high frequency. Selection of transformants was based on the expression of kanamycin resistant gene, -glucuronidase (GUS), nopaline synthase and hybridization of chromosomal DNA with GUS probe labelled with digoxigenin-ll-dUTP. The optimal conditions for transformation depended on the type of tissue, the concentration of A. tumefaciens and the use of feeder layer. Tissues collected from 20-65 days old papaya seedlings were suitable for transformation. Hypocotyl, stem and shoot tip were suitable for plantlet induction when cultured on MS medium containing 0.2 mg/l NAA and 0.5 mg/l BA. The concentrations of A. tumefaciens of 5.5*10** (8)-1*10** (9) cfu/ml were shown to be very effective. Incubation of cultured tissues with A. tumefaciens and the feeder layer in the dark for 2 days followed by the selection of transformants on the medium containing 100 mg/l kanamycin, 500 mg/l carbenicillin and 100 mg/l cefotaxime resulted in a high frequency of transformation.
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