小麦成熟胚再生体系及基因枪转化的初步研究

以小麦品种晋麦2148的成熟胚为外植体,消毒后用解剖刀刨碎,接种于MS+VB5+11.2g·L-1葡萄糖+2mg·L-12,4D+8g·L-1琼脂糖的培养基中诱导愈伤组织,将诱导3周的成熟胚愈伤组织接种到胚状体成熟培养基(MS+VB5+11.2g·L-1葡萄糖+0.2mg·L-1IAA+8g·L-1琼脂糖)中培养4-8周,然后转接到再生培养基(1/2MS+VB5+30g·L-1蔗糖+8g·L-1琼脂糖)中进行再生成苗.接种1200块成熟胚,得到1123块愈伤组织,最终形成258株再生苗,再生率为21.5%.在建立了再生体系的基础上,用基因枪介导法将GUS基因导入成熟胚愈伤组织,Xgluc染色表明,GUS基因已经在小麦成熟胚愈伤组织中表达.将100块愈伤组织用Xgluc染色,29块愈伤组织出现肉眼可见的蓝色小点.

A preliminary study of regeneration system of wheat mature embryo and its particle biolistic transformation

Mature e mbryos of a common wheat variety Jinmai 2148 was used as explants t o induce regeneration and transformation, respectively. The embryos were isol ated from surface-sterilized mature caryopses, cut into pieces, and fil ter ed through a sterile nylon mesh, then inoculated on the MS medium with 2 mg·L ~-1 2,4 -dichlorophenoxyacetic acid to initiate calli. After 3 weeks, the embryogenic calli were transferred into the MS medium with 0.2 mg·L~-1 IAA an d cultured for 4-8 w eeks for plant regeneration. Following the above protocol, 1123 calli and 258 plantlets were obtained from 1200 embryos in total,and the regeneration rate wa s 2 1.5% Furthermore,plasmid DNA of pAHC25 containing GUS gene as the d onor of alie n gene was transformed into the mature embryo calli by bombardment. The result from X-gluc staining showed that GUS gene gave transient expression success full y in the transformed calli. Blue spots were observed in 29 calli out of 100 cul tures.