Study On Agrobacterium-mediated Genetic Transformation Of HbNHX₁ In Populus Pseudo-simonii Kitag

Poplars belong to Salicaceae Populus, whose genomes are small. They are the natural hosts of agrobacterium, so they become the model species of forest genetic improvement research. Populus pseudo-simonii Kitag's trunk is very straight. It can improve ecological environment, adjust climate, provide lumber and so on. P. pseudo-simonii .is the natural host of Agrobacterium tumefaciens and its genome is small. It widely distributed in the Northeast Plain region. P. pseudo-simonii. is easy to reproduce and highly tolerance to enviroment changes. It is honored as"tobacco"in the forest species. P. pseudo-simonii ;can grow well in the pH7.5⁸.5 soil condition. When the pH of soi is over 8.5, its growth condition will be reduced. There are many saline-alkali soil in west region of Jilin Province. For improving the salt-alkali soil enviornment and enhancing its utilization, a gene (HbNHX1, originated from Hordeum brevisubulatum) which responsible for Na+/H+ reverse transmission is transformed to P. pseudo-simonii by Agrobacterium EHA105 mediated transformation. .This research provides the theoretical and experimental foundation for the saline-alkali tolerance breeding of P. pseudo-simonii.. The results of this research are as follow:1. A fast regeneration system is build up with P. pseudo-simonii stem for explant. MS base supplemented with 0.2 mg·L^-1 6-BA and 3 mg·L^-1 NAA is the optimum callus inducing and shoot regenerating media. The best media for root formation is 1/2 MS. P. pseudo-simonii reproduction system can be effectively established by using these media, that lays the foundation for the P. pseudo-simonii Kitag 's transformation.2 A P. pseudo-simonii transformation system is established by using shoot as explant in Agrobacterium-mediated transformation. The concentration of antibiotic screening is Kanamycin 50 mg·L^-1 for callus, and 25 mg·L^-1 for root regeneration..3. NaCl resistant test is done with the transgenic and non-transgenic regeneration shoots. The root regeneration frequency of transgenic regeneration shoots in medium with 4 g·L^-1NaCl is 33%, but the non-transgenic regeneration shoots don't take root at the same salt concentration. Through the NaCl tolerance and Kanamycin resistance examination, we obtain 25 transgenic plants. The identification of gene transformation will be done in the future.4. We bring forward the method to prevent vitrification of in vitro culture and discusse the factors of in vitro vitrification.