Embryo transformation, a practical approach for realizing transgenic plants of Safflower (Carthamus tinctorius L.)

Abstract

Gene transfer system that ensured recovery of whole plant transformants was developed for safflower (Carthamus tinctorius L.). Embryo axes of germinating seeds with one of the cotyledons removed were pricked with a sterile sewing needle at the cotyledonary node and infected by gentle agitation for 10 min in a suspension ofAgrobacterium tumefaciens . Following a 24 h co-cultivation and decontamination with cefotaxime for 1 h, they were placed on soilrite moistened with water to allow germination to progress. Later, the seedlings were transferred to soil in pots where they grew into normal healthy plants in the greenhouse. The histochemical assay of an uid A gene that expresses only in plant tissues and PCR amplification of uid A and npt II marker genes were used for early determination of putative transformants, whereas Southern analysis of T0and T1plant DNA was used to confirm integration of the transgenes. The combined results indicated that the frequency of transformation was 5.3% in safflower 'A-1' and 1.3% in 'A-300'. Four T₀plants of 'A-1' yielded transformed T₁progeny. The strategy, in principle, should be applicable to all cultivars and genotypes of safflower which are susceptible to Agrobacterium tumefaciens infection. Thus far, this is the only procedure available for safflower that could successfully be used to generate whole plant transformants.