Study On Multiplication And Browning In The Callus Culture Of Cephalotaxus Mannii Hk. F.

Plant cell culture technology was used to research Cephalotaxus mannii Hk. f. callus induction, culture and browning resistance in this paper, further for suspension cell culture system and the establishment of harringtonine regulation of secondary metabolism to provide experimental basis.In order to reduce contamination rate of Cephalotaxus mannii Hk. f. in the process of plant tissue culture, the branches and leaves of Cephalotaxus mannii Hk. f. were used as explants for callus induction, mercuric chloride, sodium hypochlorite and ozone were used as the disinfectants. The impact of different sterilization methods on contamination rate and cell activity of explants was studied in this paper. The results were as follows:The combinative sterilization of mercuric chloride and ozone showed the best effect. Sterilization conditions were as follows:75%alcohol for1min, ozone for30min and0.1%HgCl2solution for10min. The sterilization strategy resulted in5%of contamination rate of explants and95%of callus induction rate, enhanced38%of cell activity than the control under the sterilization conditions indicated as above.fresh leaves were the most suitable material for explants.Effects of different kinds and concentrations of antibrowning agents and hormones, as well as explants of Cephalotaxus mannii Hk. F. to the development and browning of the callus, were studied in this paper. Cephalotaxus mannii Hk. f. was used as the material, the growth rate, PPO activity, POD activity and polyphenol content of the callus were determined to assess the effects. Results were as follows:the medium added0.5g-L"1PVP could help callus obtain the maximum growth rate and effective antibrowning. The browning rate of callus induced from the leaves was lower than the one from the stems.The best proliferation medium for Cephalotaxus mannii Hk. f. was as follows:MS+4mg-L-1NAA+0.1mg-L-1KT+30g·L-1sucrose+6g-L-1carrageenan+0.5g-L-1PVP.The testing conditions of high performance liquid chromatography were as follows: mobile phase0.02mol/L; ammonium acetate:methanol=55:45; column temperature25℃; detection wavelength288mm; sample quantity lOμL; the flow rate1.000mL/min. it was proved that extracts of Cephalotaxus mannii Hk. f. contain harringtonine and homoharringtonine by this testing.