Study On Browning In The Tissue Culture Of Cattleya

Cattleya，hailed as“Orchids King”，is a kind of famous ornamental flower in the world.The paper based on the big flower variety Rlc.Beauty Girl‘KOVA’as the experimentalmaterial,its buds on the robust plants were taken as the explants,carried out the study ofCattleya browning in the process of tissue culture,explored the suitable method of Cattleyaexplants in the primary culture and subculture by adjusting the composition of the medium andculture conditions,through the comprehensive analysis of various factors,won the best mediumand culture conditions to prevent browning,and on that basis,improved the survival rate ofCattleya in the tissue culture,thus optimized the callus induction and proliferation system.Themain results were as follows:（1）With temperature of25±2℃light intensityof1500－20001x, illumination time of12h/d, pH5.5as the basic culture conditions, carried out the study of Cattleya browning in theprocess of callus induction, the results showed that medium composition, dark culture,absorbent and antioxidant had great influence on explants browning.Using medium、sucrose and agar for orthogonal experiment,the optimal medium were1/2MS+sugar20g/L+agar0g/L, at this time,the browning rate was46.7%. on this condition,cultured explants at different times of darkness, the results showed that dark culture at differenttimes could delay the process of explants browning,with extension of dark culture time,thebrowning rate decreased,but if the time was too long, it would reduce the survival rate ofexplants, this study suggested that10days of darkness after inoculation worked the best, thebrowning rate was36.7%.On the basis of the best mediun and dark culture time,added different kinds andconcentrations of adsorbent and antioxidant, the results showed that:Different kinds and concentrations of adsorbents had different effects to the browningdegree,in this experiment,added activated carbon can reduce the browning of explants,the best concentration was0.5g/L(the browning rate was13.3%),if the concentration was too high, itwould aggravate the browning degree,while the effects of PVP were better than activatedcarbon,and the concentration of6g/L worked the best,the browning rate was only6.7%.Three kinds of antioxidants (sodium thiosulfate, ascorbic acid and citric acid) were used inthis experiment,All of them postponed the time of browning,of which500mg/L of citric acidwas the best,the effects of sodium thiosulfate was the worst,while the use of VC should becontrolled within a certain range,if the concentration was too high, it would aggravatebrowning.In conclusion, in Cattleya callus induction,added1/2MS+sugar20g/L+agar0g/L+PVP6g/L,selected10days as dark culture time worked the best,and the best absorbent PVPhad a better effect than the best antioxidant citric acid.(2) With temperature of25±2℃, light intensityof1500－20001x, illumination time of12h/d, pH5.5as the basic culture conditions, carried out the study of Cattleya browning in theprocess of callus proliferation, the results showed that different days of subculture, plantgrowth regulators and adsorbent had great influence on callus browning.In Cattleya callus proliferation,the suitable saaubculture cycle to prevent browning was21days,at this time, the callus proliferation multiple was3.237, the browning rate was26.7%.Onthis basis selected the suitable hormone combination was3mg/L6-BA and0.5mg/LNAA,itcan not only improve the callus proliferation multiple(3.333), but also can greatly reduce thecallus browning rate(20%).Under these conditions added different adsorbent can not only reduce the browningrate,but also can ensure the callus proliferation,the effects of PVP were better than that ofactivated carbon,it was necessary to control the concentration of activated carbon in thepractical application,in this study,3g/LPVP worked the best to the callus culture, the callusproliferation multiple was3.519, the browning rate was13.3%.In conclusion, in Cattleya callus proliferation,added3mg/L6-BA+0.5mg/LNAA+3g/LPVP,selected21days as the subculture cycle worked the best.