独行菜eEF-1a基因片段分离克隆及RT-PCR分析

Cloning and RT-PCR Analysis of eEF-1a Gene Fragment from Lepidium (Lepidium apetalum Willd.)

文章根据已知植物eEF-1a(编码Eukaryotic elongation factor1-alpha)基因的保守序列设计引物,采用RT-PCR的方法扩增独行菜(Lepidium apetalumWilld.)eEF-1a基因片段。结果获得一大小为570bp的基因片段,序列比对表明,该基因片段与拟南芥eEF-1a基因核苷酸序列的同源性达到95%,推测该其应该是独行菜eEF-1a基因片段。采用RT-PCR方法,对独行菜三个生长阶段材料及对应阶段冷诱导后地材料中的eEF-1a基因表达作了分析,表明eEF-1a在独行菜组织中表达稳定,可以作为实时荧光定量PCR研究独行菜基因表达分析中的看家基因。

Degenerate primers were designed based on the conserved sequences of the eEF-1a genes from other plants. And the eEF-1a gene fragment was obtained by RT- PCR from Lepidium （Lepidium apetalum Willd.）. The eEF-1a gene fragment contained 570 bp nucleotide acids. Blasting analysis showed that it has 95% nucleotide sequence homology with one in arabidopsis thaliana. And RT-PCR analysis showed that the expression of eEF-1a gene was steady in Lepidium, and it can be used as internal standard when study on other genes’ expression and regulation in Lepidium.