高羊茅甘油醛-3-磷酸脱氢酶基因FaGAPDH的克隆与分析

Cloning and Characterization of Glyceraldehyde-3-Phosphate Dehydrogenase Gene FaGAPDH in Tall Fescue

以高羊茅茎基部与叶基部的mRNA为模板,引用基于多年生黑麦草GAPDH基因序列设计的引物,进行RT-PCR分析,同时结合3'RACE和5'RACE方法从高羊茅中扩增出两个GAPDH基因,命名为Fa-GAPDH1(GQ480772)与FaGAPDH2(GQ480773)。两序列cDNA全长分别为1281bp和1348bp,具有完整的开放阅读框(ORF,FaGAPDH1:74～1087bp;FaGAPDH2:106～1119bp),编码蛋白都为337个氨基酸。保守结构域功能分析表明两基因编码的蛋白质都具有典型的Rossman-NAD连接折叠和C-末端结构域。FaGAPDH1和FaGAPDH2蛋白与禾本科植物的该基因蛋白产物比较发现氨基酸序列的同源性都在90%以上,说明两基因具有高度的保守性。

Taking the stem base and the leaf base of tall fescue as temples, with primers designed based on the sequence of GAPDH gene from Lolium perenne L., two full-length cDNAs of GAPDH genes were cloned from tall rescue by RT-PCR combined with 3＇RACE and 5＇RACE. The two newly cloned GA PDH genes were designated as FaGAPDH1 （GQ480772） and FaGAPDH2 （GQ480773）. Sequence analysis shows that FaGAPDH1 is 1 281 bp in length, and contains a single open reading frame from 74 bp to 1 087 bp. FaGAPDH2 is 1 348 bp in length, and contains a single open reading frame from 106 bp to 1 119 bp. They both encode proteins with 337 amino acids. Domain analysis reveals that proteins encoded by the two genes both contain two conserved domains： NAD（P）-binding Rossmann-fold domains and C-terminal domain. Similarity comparison between FaGAPDH1 protein, FaGAPDH2 protein and the protein encoded by GA PDH gene from gramineous plants shows that they own amino acids sequence similarities of more than 90%, indicating that the two new genes are both highly conservative.