影响草地早熟禾(Poa pratensis L.)基因枪转化的关键因素研究

Study on the Bombardment Factors for Kentucky Bluegrasss (Poa pratensis L.)

将含有DREB1A基因的表达载体,通过基因枪法转化草地早熟禾的胚性愈伤组织,探讨了金粉沉淀剂、金粉直径等因素对转化的影响,同时通过不同浓度潮霉素(Hygromycin,简称Hy)对未转化愈伤组织的筛选,获得最佳筛选浓度。结果表明,适合于草地早熟禾基因枪轰击的条件为Ca(NO3)2+PEG4000包被质粒DNA;1μm金粉作为质粒DNA的载体;合适的轰击高度为6cm、轰击次数为1次、无渗透处理。采用Hy作为草地早熟禾转基因植株抗生素筛选标记时,Baron品种愈伤组织继代的临界筛选浓度为100mg/L。

Embryonic callus of Kentucky bluegrass were used as material for genetic transformation with the DREB1A gene by particle bombardment. Parameters of biolistic bombardment were studied. The optimal methods showed as follows： plasmid DNA are coated by Ca（N03）2 and PEG4000, 1μm golds are the vectors of plasmid DNA, 6 cm bombardment height distance, 1 time and without osmotic treatment are favorable to transgenic efficiency. The concentration hygromycin （Hy） , which is the selection mark for cuhivar＇ Baron＇ is 100mg/L.