Study On CBF3 Of Arabidopsis Thaliana Transfered Into Hevea Brasiliensis

The CBF3 gene which controls the plant’s cold resistance was transferred into the callus of Hevea brasiliensis Reyan 7-337-97 by Particle Bombardment and Agrobacterium-mediated method separately.In the Aagrobacterium-mediated gene transformation method, some factors which could impact the transgenic rate such as the acceptor materials, the co-culture time after infection, the co-culture temperature and the concentration of antibacterial antibiotics were studied. The result showed that the callus could effectively resist the Agrobacterium contamination and they could continue to grow under the conditions that:the anther callus was cultured for 30 days before being infected, being infected for 3min, co-cultured for 2 days after the infection in 20℃on the medium with Cef 200mg/L. The rate of embryo differentiation from anther callus is very low and the Cef for bacterium inhibition is harmful to the callus. So it was difficult to generate normal embryo from the being infected callus.In the Particle Bombardment method three resistant seedlings were obtained through the 25mg/L Hyg resistance selection. In the resistant seedlings’ leaf DNAPCR detection, one was positive. This transgenic plantlet was propagated for 4 generations and 29 new seedlings were obtained. PCR of all the extracted leaf DNA were positive. This result showed that the CBF3 gene probably had been integrated into the genome of rubber preliminary. BamHI and HindⅢwere used to digest the DNA separately. Then the result of Southern Blot identification showed a clear signal in the transformed plantlets while there was no signal in the non-transformed plantlets’ genomic DNA. It demonstrated that the CBF3 gene had been integrated into the genome of Hevea brasiliensis Reyan 7-337-97 successfully and its’ genetic stability. Some of the propagated transgenic plantlets would be transplanted into field soon.