蒺藜苜蓿DGAT1基因的克隆和功能鉴定

该研究采用RT-PCR与电子克隆的方法,从蒺藜苜蓿cDNA中克隆得到2个编码二脂酰甘油酰基转移酶(diacylglycerol acyltransferase,DGAT)的基因MtDGAT1-1和MtDGAT1-2。MtDGAT1-1长1 620bp,编码539个氨基酸;MtDGAT1-2长1 524bp,编码507个氨基酸。多序列比对显示,MtDGAT1-1和MtDGAT1-2编码蛋白具有典型的植物DGAT1结构域。表达分析显示,MtDGAT1-1和MtDGAT1-2在根、茎、叶、花、种子中都有表达,在种子发育中高表达,且MtDGAT1-1于种子发育的中前期高表达,而MtDGAT1-2于种子发育的中后期高表达。酵母互补实验证实,MtDGAT1-2编码蛋白具有DGAT酶活性,能够恢复H1246的TAG合成和油体形成;而MtDGAT1-1编码蛋白不能恢复H1246的TAG合成和油体形成。

Cloning and Functional Characterization of Diacylglycerol Acyltransferase Gene (MtDGAT1) from Medicago truncatula

In this study, we acquired two MtDGAT1 genes (MtDGAT1 1 and MtDGAT1 2) from cDNA of Medicago truncatula by using insilico cloning combined with RT PCR. The length of MtDGAT1 1 is 1 620 bp, encoding 539 amino acids, while the length of MtDGAT1 2 is 1 524 bp, encoding 507 amino acids. Multiple alignments revealed that both MtDGAT1 1 and MtDGAT1 2 shared the typical functional motifs with DGAT1s from other plant species. Expression analysis showed that both MtDGAT1 1 and MtDGAT1 2 were expressed in root, stem, leaf, flower and seed, and highly expressed in developing seeds; MtDGAT1 1 was highly expressed in early middle periods of seed development, while MtDGAT1 2 was highly expressed in the middle and later periods of seed development. Functional complementation in yeast H1246 confirmed that MtDGAT1 1 encodes disfunctional DGAT and MtDGAT1 2 encodes functional DGAT.