| Phellodendron amurense Rupr. has important economical and medicinal value. The conditionof P. arnurense Rupr. seeding request to be high, the technical difficulty is big, the transplantsurvival rate is low, slow-growing, the age limit to be a useful adult is long, so it is notpromoted by cultivation in big area, the wild resources lack, and now exist these problem suchas natural reproduction is difficult, the bough shape is not straight, planted forest seed abortand so on. This research considers existing question of P. amurense Rupr., uses P. amurenseRupr. seed directly to carry the tissue culture for thefirst time, establishes P. amurense Rupr.fast and effective cultivate system, achieves heredity transformation for P. amurense Rupr. forthe first time, establishes fast and effective transformation gene system, establishes thefoundation for the development of P. amurense Rupr. improved seed.1. The research of tissue culture of P. arnurense Rupr. indicates:The comeback-injure organization of Huang Bo's seed under the function of 6-BA and NAAoccur most early, the rate of P. amurense Rupr. abduction in the substrate with appending 4.0mg·L-1 TDZ和0.2 mg·L-1 NAA under the condition of 28℃, the average number of bud byabduction is the most. The basic substrate's allocated proportion in favor of the growth of P.amurense Rupr. bud is the 1/2 MS substrate appended 2 % cane sugar.In the process of bud growth, the consistency of 6-BA plays the pivotal effect, NAA has theeffect of promoting bud growth. The substrate with appending 4.0 mg·L-1 6-BA and 0.2 mg·L-1NAA is best for bud growth, and the completely growth quantity reaches 3.75 cm in twoweeks.The radical revulsive rate of bud in medium adding 4.0 mg·L-1 6-BA and 0.2 mg·L-1 NAAachieved 85 %. Low consistency of 2,4-D made for inducing embryo genesis of P. amurenseRupr, it can induce embryo genesis in the medium appending 1.0 mg·L-1 2,4-D or in themedium appending 4.0 mg·L-1 TDZ and 0.2 mg·L-1 NAA, the rate of inducing was 16.67 %,1.22 %, 32.26 % and 2.02 %. The endosperm of P. amurense Rupr. induced a bud in mediumadded 4.0 mg·L-1 BA and 0.2 mg·L-1 NAA.2. The research about astablishment of agrobacterium-mediated transformation of P. amurenseRupr. Showed:The method that cut every seed to two half in germ liquid was advantageous to theagrobacterium to combine with seeds and it could obtain high efficiency of fastness bud. Theliquid medium to dilute agrobacterium is 4.0 mg·L-1 TDZ+0.2 mg·L-1 NAA. Inducing mediumof fastness bud add 200 mg·L-1 cefotaximem and 10 mg·L-1 kanamycin. The infected seedswere co-cultivated with agrobacterium for 2 days in 28℃and 2 days in 4℃in darkness. Theendosperm should be saved during seeds were co-cultivated with agrobacterium, and be peeled off after that. The callus should be cut into 0.5cm×0.5 cm×0.5 cm during the fastness budproliferate and develop. The livability of the transgenic P. amurense which was cultivated inwater for 3 days achieved 100%. In this experiment, we obtained 70 transformants, thenrandom chose 5 transformants to check. The result indicated that the extrinsic gene LEA hadinserted into genome of accepter by PCR amplification, PCR-Southem hybridization and RT-PCR amplification.
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