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以大麦花培基因型花30的幼胚为外植体,设置不同的培养基类型、不同激素配比及碳源,研究其对幼胚愈伤组织诱导及绿苗分化的影响,以此建立和优化一个适于优良大麦品种遗传转化的高效组织培养体系。结果表明:在N6、MS和B5的组合改良培养基下,以蔗糖为碳源,附加2mg/L 2,4-D、1mg/L ABA时,有最高的愈伤组织诱导率,且愈伤质量最好。Cu2+的添加具有抑制幼胚直接发芽成苗和改善愈伤组织质量的双重功效。添加2mg/L 6-BA对愈伤组织的分化效果比较理想。为了提高农杆菌介导转化大麦外源基因的瞬时表达率和优化遗传转化体系,利用花30幼胚产生的愈伤组织为受体材料,通过检测GUS基因的瞬时表达情况,研究了农杆菌介导的大麦遗传转化中菌液的浓度、侵染时间以及共培养天数对遗传转化的影响,结果表明:当菌液浓度OD600=0.5的条件下,侵染15min,共培养2d表现出最佳的GUS瞬时表达率。
Using the immature embryo of genotype flower 30 of barley as explants, the effects of different medium types, different hormone ratios and carbon sources on the induction of embryo callus and the differentiation of green plantlets were studied. And optimize an efficient tissue culture system suitable for genetic transformation of elite barley varieties. The results showed that under the combination of N6, MS and B5, the highest callus induction rate was obtained with sucrose as carbon source, 2mg / L 2,4-D and 1mg / L ABA, and the callus The best quality. The addition of Cu2 + has the dual effect of inhibiting the germination of immature embryos directly and improving callus quality. The effect of adding 2mg / L 6-BA on callus differentiation was ideal. In order to improve the transient expression rate of the foreign gene mediated by Agrobacterium tumefaciens and optimize the genetic transformation system, the callus generated from the embryos of flower 30 was used as the receptor material. By detecting the transient expression of GUS gene, The results showed that when the bacterial concentration of OD600 = 0.5, the infection 15min, co-culture 2d showed the best GUS transient expression rate.