红三叶新品系无菌苗培养体系优化

Optimization for Aseptic Culture of Red Clover(Trifolium pratense L.)

以红三叶(Trifolium pratense L.)新品系为材料,对无菌苗培养的种子消毒方法和培养基组成进行研究,以期为红三叶新品系组培再生体系、分于育种及抗病性研究提供基础依据。结果表明:0.1%HgCl2,3%NaClO和15%H2O2消毒不超过15 min,对红三叶新品系种子发芽率没有显著影响,但消毒效果存在较大差异。红三叶种子消毒的最佳方法为75%酒精预处理,0.1%HgCl2消毒10 min,发芽率可达89.44%,并彻底无污染。最适培养基为1/4MS+1.5%蔗糖,不添加激素,pH6.0,发芽率可达85.78%,无菌苗生长良好。培养基中无机盐的含量是影响红三叶种子发芽率和无菌苗生长的关键因素。琼脂培养基培养比滤纸培养的无菌苗子叶和下胚轴出愈率显著提高,但不同组分琼脂培养基的无菌苗出愈率无显著差异。

Effects for both seed disinfection methods and medium composition on tissue culture were investigated in red clover（Trifolium pratense L.）.Results show that seed germination rate has no significant differences using 0.1% HgCl2,3% NaClO,or 15% H2O2 sterilization for less than 15 min.However,the efficiency of sterilization is different between the three disinfectants.The best method of seeds sterilization for red clover is the 75% ethyl alcohol treatment followed by disinfection with 0.1% HgCl2 for 10 min.Germination rate reaches 89.44% and is completely pollution-free.The most suitable medium is 1/4MS plus 1.5% sucrose without hormones at pH 6.0 for seed culture.Germination rate reached 85.78% and seedlings grew well with this medium.The content of inorganic salts in medium is one of the key factors affecting seed germination and growth in aseptic cultures.Callus induction rates of cotyledons and hypocotyl are significantly increased in agar medium as compared with filter paper,but with no significant difference among different components agar medium.