Studies On Tissue Culture And Means On Based Hairy Roots System Of Saposhnikovia Divaricata

Saposhnikovia divaricata (Turcz.)Schischk. and Agrobacterium rhizogenes were taken as the material in this dissertation. Acquired antiseptic seeding of Saposhnikovia divaricata (Turcz.)Schischk, used orthogonal test plan,chose its best of all substrate in apiece phase of tissue culture. It studied the way on made the callus culture system and its aftergrowth of Saposhnikovia divaricata (Turcz.)Schischk. And, it studied Agrobacterium rhizogenes was appraised used physiology biochemistry and molecular biology, biology detected and energy compared. And, Saposhnikovia divaricata (Turcz.)Schischk.'s genetic transformation with Agrobacterium rhizogenes R1000 were investigated as well as changes in the contents of sucrose, its hairy root cultures system was made. The main results were as follows:It was used paper bridge means(1/2MS) when based its asepsis system, these seed were polluted percent could decline 13.3% as planting seed capsule shuck of, and come up out of land change 7d, burgeon percent is 65.2%, its germination percentage achieved 65.2% (A mature seed germination percentage is 50% at nature); It is obvious that the callus as 7d its laminae explant in MS+2,4-D0.6+6-Ba0.5 substrate, The callus come into being percent is 100%; there into caudex segment and root's explant recover percentage were high and cubage,eugonic that the best substrate used culture the callus, it is MS+6-BA1.0+NAA0.6,the callus's proliferate effect achieve best salience level in contain 6-BA,NAA substrate and the callus's proliferate effect not salience in contain KT substrate. its proliferate multiple achieve 13.68 double on 30d; that the best substrate used bud polarization of culture the callus, it is MS +NAA0.6+KT0.5+6-BA1.0,its bud polarization percent achieve 70%; its adventitious bud procreant root is white and thickset in MS+NAA0.4 substrate, its rhizogenesis percent is 100%, its seedling could transplant when its root came 3~4cm, its seeding high about 5cm.There were seven bacterium were appraised used physiology biochemistry, thereinto R1025,R1601,R10001 were biologyâ…¡model Agrobacterium rhizogenes, it is identical as a result with PCR appraised (R10001,R1025,R1601 have evidence brigh at thereabout 540bp,770 bp). The callus was cultured for 15h in the liquid YEB substrate, the bacterium density leave each other is R1601>R10001 >LBA9402>ATCC15834>R1025>R10002>A4. It determined provide test bacterium genetic transformation percent using cucumber's young cotyledon, The result showed that R10001's genetic transformation percent is most, it is 59.0%, every explant average rootage is 4.21 item. The Cef 400mg/L's effect was best on antibiotic was chosed.The young cotyledon is inducement percent highest at based the hairy root system process. Three expiant were inducement percent devolution were: cotyledon>caudex>root. The young cotyledon was cultured 2d, it is inducement percent is 34.29% then nought Treatment; They common culture best time is 2d, If time prolong that Agrobacterium rhizogenes was a great deal reproduced, further more explant will perished; At preparation culture, AS best append quantity is 30 mg/L, the effect wasn't salience at add dosage;Use ultrasonic Treatment effect was preferably front contagion, but it must was short time, Or else conduce explant plentiful death; Saposhnikovia divaricata (Turcz.)Schischk.'s cotyledon was inducement best term : beforehand substrate adding AS30mg/L, Use ultrasonic Treatment front contagion 4s, after ,together culture 2d, its inducement percent is 62.9%. The curve develop hairy root of Saposhnikovia divaricata (Turcz.)Schischk. was mensurated in liquid MS substrate. It was soon in 5~20d, along with culture time protract to 35~40d, its hairy root growth entered roof garden period, slowness growth. After 35d, its basal hairy root was graveness browned.Saposhnikovia divaricata (Turcz.)Schischk. fair roots could come into being callus onto MS+2,4-D0.6+6-BA0.5 substrate,,Adventitious buds can be observed to form gradually from the calli on MS medium supplemented with NAA0.6mg/L, KT0.5 mg/L and 6-BA1.0 mg/L.