陆地棉GhDGAT1基因干涉载体构建与遗传转化

Construction and transformation of RNAi vector of gene GhDGAT1 in upland cotton

二酰基甘油酰基转移酶(DGAT)是生物体内三酰甘油(TAG)合成过程中的关键酶。本研究克隆陆地棉GhDGAT1基因308bp片段,构建了该基因含内含子结构的hpRNA干涉载体;应用花粉管通道法转化棉花,研究内源基因GhDGAT1沉默对棉花油分含量的影响。结果表明:1)经PCR及Southern杂交鉴定,转基因种子中GhDGAT1基因表达量受到显著抑制,种仁含油量下降,最多下降3.13%。2)种子油脂量显著减少的株系中,总蛋白含量及可溶性糖分含量,分别相对提高4.31%～9.77%和11.67%～23.01%。3)与野生型相比,转基因株系幼胚在发育后期鲜重降低,而可溶性蛋白含量升高。4)与对照相比,转基因植株的株高、第一果枝高度、果枝数均显著降低,但其他农艺性状与主要的经济性状均未受到显著影响。研究表明,通过调控GhDGAT1基因的表达可影响陆地棉的油脂合成,调节棉籽油分含量。

A 308-bp fragment of cotton GhDGAT1 gene was used to create intron-containing construct expressing hpRNA vector,and transferred to cotton via pollen tube path way,for studying the impact on oil content by silencing the endogenous GhDGAT1 gene of cotton. Results indicated that.. 1） After PCR and Southern blotting, the expression of GhDGAT1 gene were inhibited significantly in T3 transgenic seeds,and the oil contents of seeds were decreased up to 3.13 percent. 2） In transgenic lines with lipids contents decreased, the protein contents and soluble sugar contents were relatively increased 4.31% - 9.77 % and 11.67 % - 23.01%, respectively. 3） Compare to wild type, the fresh weights of immature embryos in transgenic lines were reduced in the later stages of embryos development, but increased in soruble protein contents. 4） Compare to wild type, the plant height, height of primary branch and branch number in transgenic lines were decreased significantly,but have no significantly changes in other agronomic traits and the main economical traits. The finding that GhDGATI gene influenced the lipids synthesis in cotton could be used to regulate oil contents in kernels by modulating the expression of GhDGAT1.