低能氩离子束介导将绿色荧光蛋白基因导入小麦的研究

Study on transferring improved green fluorescent protein gene into wheat via low energy Ar^+ implantation

用低能氩离子束介导将改良的绿色荧光蛋白基因 (mGFP4)导入小麦栽培品种皖 92 10和皖麦 32号的成熟胚细胞。在含有 10 0～ 140mg/L巴龙霉素培养基上继代培养后 ,获得了一批抗性愈伤组织。经过分化培养 ,皖 92 10获得 5株再生苗 ,皖麦 32号获得 32株绿苗 ,对照的 2 0 0枚成熟胚未获得再生苗。对再生苗进行PCR检测 ,均扩增出 6 0 0bp的nptⅡ基因片段。取 4株长势好的绿苗进行Southern杂交分析 ,结果表明这 4株绿苗皆为转基因植株。根据PCR分析结果统计 ,皖 92 10和皖麦 32号的平均植株转化率分别是 1 3 %和 4 1%

An improved GFP gene(mGFP4)was introduced into mature embryo cells of wheat cultivars Wan 9210 and Wanmai 32 via low energy ion beam mediated delivery technique.Resistant calli were selected on medium containing paromomycin(100?140 mg/L).Five green plants were regenerated from resistant calli of Wan 9210 derived from 387 implanted mature embryos.32 green plants were obtained from 776 irradiated mature embryos in Wanmai 32.No green plant was regenerated from calli of 200 non transformed embryos.PCR assays of 37 green plants showed that they all obtained the expected size of amplified DNA fragment(600 bp).Southern blot of 4 well?developed green plants confirmed stable integration of GFP gene into wheat genome.The average transformation frequencies of Wan 9210 and Wanmai 32 were 1 3% and 4 1%,respectively,according to the results of PCR assays.