The Preliminary Reseach In Tissue Culture About Two Kinds Of Clematis

Clematis mandshurica and Clematis fusca are both the important wild species of Clematis.They had some ornamental characteristics and medicinal value. However, there was less research of these two Clematis now. It would be very important to use scientific and effective methods for both wild species of Clematis germplasm preservation.In this paper, the culturing stems with buds, stems and leaves from two kinds of Clematis were the explants in the tissue culture of two kind of Clematis, which would provide the basic information for the rapid promotion of wild. The results were as follows:1. The tissue culture in cluster shoots induction from the stems(1)The creation of sterile system:Stems with buds were used as explants of the two wild species of Clematis in China. The best sterilization time by HgC12(0.1%) was5minutes to the stems with buds.(2)Collecting season:Spring was the best season to collect the explaints.It had the highest germination rate and lower pollution rate of the explants.(3)Germination of stems with buds:The best initial medium of Clematis fusca was MS+6-BA1.0mg·L-1+NAA0.05mg·L-1,The germination rate was93.33%. The best initial medium of Clematis mandshurica was MS+6-BA2.0mg·L-1+NAA0.01mg·L-1The germination rate was86.67%.(4) Proliferation from stems with buds:The best culture medium of proliferation from stems with buds on Clematis fusca was MS+6-BA2.0mg·L-1+NAA0.05mg·L-1+KT1.0mg·L-1, the proliferation index of adventitious buds was2.63. The best culture medium of proliferation from stems with buds on Clematis mandshurica was MS+6-BA1.0mg·L-1+NAA0.5mg-L"1, the proliferation index of adventitious buds was3.94.(5) Seedlings and rooting culture:Through this experiment,MS and1/2MS without any hormones had failed to meet two Clematis’buds seedlings results and had failed to found the rooting medium.2. Preliminary study of callus induction on the stems and leaves(1)The creation of sterile system:Stems and leaves were used as explants of the two Clematis. The best sterilization time by HgCl2(0.1%)was5minutes to the stems. The best sterilization time by HgC12(0.1%)was3minutes to the leaves.(2) Effect of different hormone combinations on the callus induction:The best culture medium of callus induction from stems on Clematis fusca was MS+6-BA1.0mg·L-1+NAA0.5mg-L"1, induction rate was96.42%. The best culture medium of callus induction from stems on Clematis mandshurica was MS+6-BA1.0mg-L"’+NAA0.1mg-L’1, induction rate was80.77%. (3)Effect of different hormone combinations on the callus proliferation and differentiation: The best culture medium of callus proliferation on Clematis fusca was MS+6-BA2.0mg-L"’+NAA0.5mg·L-1, proliferation rate was52.17%. The best culture medium of callus differentiation on Clematis fusca was MS+6-BA2.0mg·L-1+NAA0.05mg·L-1. The best culture medium of callus proliferation on Clematis mandshurica was MS+6-BA1.0mg·L-1+NAA0.02mg·L-1, proliferation rate was60.87%. The best culture medium of callus differentiation on Clematis mandshurica was MS+6-BA2.0mg·L-1+NAA0.02mg·L-1.(4)Effect of different levels of activated carbon(AC) and subculture cycle on callus browning:When activated carbon on the levels of2.0g·L-1and the subculture cycle was20days, the lowest browning rate would be get.(5)Effect of light on the callus induction:The light should be better for the callus induction of leaf on Clematis fusca.3. The change of physiology-biochemical indexes on callus of Clematis mandshuricaThe growth-curve of callus looked like "S" in the light and dark.The growth-circle could be divided into3parts:period of delay(0-9d), period of growth(9-21d), period of stability (21-27d).The changes of POD actives, SOD actives, soluble protein contents and soluble sugar contents were related to the growth-curve and was related to light and dark.