Abstract:

This paper reports a successful transformation of chitinase gene into Populus simonii×P. nigra by Agrobacteriuma-mediated means. For reducing adventitious buds, the optimal concentration of kanamycin was 40 mg·L^-1. Up to 700 mg·L^-1 of Cefazolin Sodium had no obvious effect on differentiation of leaves. About 20-day-old leaf disc explants were pre-cultured for 3-4 d, then immersed in Agrobacteriuma suspension for infection for 6-15 min, then co-cultured on non-selection culture medium for 3 d in the dark at 25 ℃. At last, the explants were transferred to the selection culture medium (containing kanamycin 40 mg·L^-1 and Cefazolin Sodium 700 mg·L^-1) in light at 25 ℃. RT-PCR analysis showed that 8 of 13 transgenic plants, which were screened by PCR and PCR-Southern blot analyses, had target bands. This result confirmed that the target gene had been integrated into genome of Populus simonii×P. nigra and expressed stably on the level of transcription. Chitinase activity of most transgenic plant leaves was higher than that of non-transgenic plants obviously, and No.11 transgenic plant had 3.093 times chitinase activity of the control plant.

Key words: Populus simonii×, P. nigra, transformation, chitinase gene, chitinase activity