杨梅酸性转化酶基因cDNA分离及表达分析

Isolation and expression analysis of acid invertase gene cDNA from Myrica rubra

杨梅果实富含蔗糖,酸性转化酶是蔗糖代谢关键酶,根据植物酸性转化酶基因保守区序列设计引物,提取杨梅叶片RNA,逆转录获得cDNA,以此为模板通过PCR技术扩增到长度为516bp的基因片段,克隆入pMD18-T载体中,命名为MrIVR1(GenBank:DQ339699)。测序及同源性检索表明,该基因推导氨基酸序列与君子兰、葡萄、草莓、胡萝卜等酸性转化酶基因氨基酸序列同源性为60%～69%。运用ClustalX软件对植物转化酶基因进行了系统树分析,结果显示,MrIVR1编码的蛋白质属于细胞壁酸性转化酶。半定量RT-PCR表达分析显示,MrIVR1基因在杨梅果实发育早期表达量最高,随着果实的发育表达量下降,在成熟果实中表达水平较低。

An acid invertase gene was cloned from Myrica rubra Sieb. et Zucc. by using RT-PCR and designed primers of conserved domains of the plant acid invertase genes＇ nucleotide sequences. The fragment cloned named MrⅣ1 （GenBank number： DQ339699）, was 516 bp in length, encoding 171 amino acids. The amino acid sequences were 60%-69% identical to the sequences of orchid, grape, strawberry, carrot, etc. The putative protein encoded by MrⅣ1 was belonged to cell wall invertase by phylogenetic analysis of using clustlx software. Semiquantitative RT-PCR analysis showed that the MrⅣ1 gene was highly expressed in young fruit. The expression level was gradually decreased and reached the lowest level in the mature fruit.