Agrobacterium Rhizogenes-mediated Transformation Of Panax Quinquefolium

Vaccine seedings and three-year roots of P.quinquefolium are transformed by 3 types of Agrobacterium rhizogenes(R1601,A4,A4(rolA)). The test analyses 6 facters for increasing the transformation rates of P.quinquefolium. Facter â…  , types of explant of P.quinquefolium; facterâ…¡ ,types of A.rhizpgenes; facterâ…¢,density of A. rhizpgenes; facterâ…£, whether or not to have acetosyringone; facter â…¤,times of preconditioning and co-cultivation with P.quinquefolium and A.rhizpgenes; facterâ…¥,time of infection. The test still analyses the growth of hairy roots from P.quinquefolium and the control of content of secondary metabolite according as several facters of chemistry and acquires the production system of secondary metabolite according as the transformation of hairy roots from P.quinquefolium.(1) The system of hairy roots from P.quinquefoliumSeeds are cultivated in the pot, which was maintained at approximately 25 ℃ and shading duing the day. Seedlings (1²weeks) are cut into three different parts (leaves, stems and seedlings with leaf explants) as explants. The high transformation rate will be acquired when the suitable density of A.rhizpgenes(OD₆₀₀,AS=100μmol/L), preconditioning time (48h), co-cultivation time (48h) and time of transformation(20min).We tap A.rhizpgenes(R1601,A4,rolA, 28℃ illumination 120r/min) 1 ml and join As(100μmol/L) to activate 6h and smear them to the tangent plane of three-year root(25℃ unillumination). The transformation rate of R1601 and A4 is 12.5% and 21.2%, and rolA could not transform hairy root. The 100μmol/LAS could advance the transformation of hairy root.(2) Examination of hairy rootThe test examines the hairy root from seedlings with leaf by A.rhizpgenes of R1601.Hairy root from P.quinquefolium has growth of good velocity and does not need exogenous hormone in morphology. The molecule biology technique has already proved T-DNA of Ri plasmid ttansfering to the gene of P.quinquefolium because PCR detectives the prospective band(564bp).(3) The control of hairy roots and secondary metabolite to use different chemical factersHairy roots are inoculated 5 kinds of culture medium and different pH of liquid culture medium to observe the influence of growth of hairy root and secondary metabolite. And hairy roots are inoculated 5 kinds of carbon source and 6 kinds of exogenous hormone to observe the influence of growth of hairy root and secondary metabolite. In result, chemical facters have an immense influence to growth of hairy...