基因枪法获得优质HMW亚基基因表达的转基因小麦

Transformation and Expression of High Molecular Weight Glutenin Subunit Genes in Common Wheat (Triticum aestivum L.) Varieties

为探讨利用基因工程技术进行小麦品质改良的可行性,用基因枪法对湖北省3个小麦品种鄂恩1号、鄂麦11号和鄂麦12号分别进行优质高分子量麦谷蛋白亚基(HMW-GS)基因1Ax1和1Dx5+1Dy10的转导。试验结果表明,经基因枪轰击的幼胚外植体的再生频率和转化频率明显比幼穗高;基因型间愈伤组织再生能力和转化频率的差异,直接影响着小麦转化的成功率;3个品种的幼胚发育时期与其再生频率均呈显著负相关(r=-0.93或-0.95);在可控环境条件下,花后12～14d的供试植株幼胚的转化频率最高(鄂恩1号、鄂麦11号和鄂麦12号分别为4.5%、2.9%和2%)。研究表明,鄂恩1号、鄂麦11号适龄期的幼胚试材,用等摩尔比的优势亚基基因1Dx5和1Dy10与选择标记基因以2:1摩尔比混合的沉淀物包裹金粉,经600～900psi氦气压轰击后,在含0.5mg·L-12,4-D的MS培养基中诱导愈伤、含0.1mg·L-12,4-D和5mg·L-1玉米素的R培养基中分化、3mg·L-1L-PPT选择压下继代分化筛选,可获得稳定可育的T0代转基因植株以及胚乳特异性表达的T1代转基因种子。

To explore the feasibility of improving wheat quality by genetic engineering, three Chinese winter common wheat varieties Een1, Emai11 and Emai12, were transformed with high-molecular-weight glutenin subunit (HWM-GS) genes 1Ax1 and 1Dx5+1Dy10 by particle bombardment, respectively. A significant negative correlation between plant regeneration frequency and the age of immature embryo were observed in all three tested varieties. The best transformation frequencies were achieved up to 4.5%, 2.9% and 2% for three varieties of Een 1, Emai 11 and Emai 12, respectively, when using immature embryos from young donor plants developing 12 to 14 days after blossoming. The phenotypically normal fertile T0 transgenic wheat plants of HMW-GS genes 1Dx5+1Dy10 and the endosperm-specific expressed T1 transgenic seeds were produced. Factors including explants at an optimal developmental stage from donor plant and optimal conditions of bombardment and tissue culture were proved to be the key parameters in the successful transformation procedure.