猪ADSL基因克隆及其在部分组织中的mRNA定量表达

试验以通城猪为研究对象,在对猪ADSL基因全长编码区cDNA片段进行克隆、序列测定、与GenBank中已收录的其他14种动物的ADSL全长编码区序列进行同源性分析,并构建系统发生树的基础上,采用实时(相对)定量PCR方法对刚出生和65日龄两个不同生长发育时期的通城猪心、肝、肾脏、背最长肌4种组织的ADSL基因表达谱进行分析。结果表明,所克隆的ADSL基因片段大小为1 548 bp(GenBank登录号:GU249574),其中包含一长为1 455 bp的完整阅读框,通过核酸序列分析发现,该基因编码484个氨基酸;与GenBank中已收录的猪ADSL基因序列同源性最高,达99.9%;与恒河猴和黑猩猩的序列同源性最低,分别为66.3%和41.3%;15种动物的20条ADSL全长CDS序列聚类分布为两个主支。刚出生时的通城猪ADSL基因在心、肝、肾脏组织的mRNA表达水平均高于65日龄通城猪的心、肝、肾脏组织中的mRNA表达;然而,其背最长肌的ADSL基因mRNA表达水平则低于65日龄的通城猪。本结果为猪ADSL基因的生物学功能研究和遗传进化研究提供了分子依据。

Molecular cloning and mRNA expression of ADSL gene in pig tissues

The full-length CDS fragment of the porcine ADSL gene was cloned,sequenced and utilized for the sequence homology analysis and the phylogenetic tree construction with those of the 14 animal species published in the GenBank in the current experiment by using a local Chinese pig,Tong Cheng pig.The ADSL gene expressions in the four tissues at two ages were examined by a relatively Real-time quantitative RT-PCR.The result showed that a 1 548 bp cDNA fragment(GenBank accession No.GU249574) was amplified and it contained a complete ORF encoding a deduced protein of 484 amino acid residues of ADSL.Analysis of the porcine ADSL sequences revealed that it was highly homologous to that of the previously published porcine ADSL CDS,99.9%,and the lowest similarity of 66.3% and 41.3% was found to that of Rhesus monkey and Pan troglodytes,respectively.The 20 sequences of the ADSL genes of 15 animal species were clustered into two main branches by the phylogenetic tree constructed using the method of Jotun-Hein.The tissue expression profile showed that the mRNA expression levels of ADSL gene in heart,liver and kidney issues at the newly born pigs were higher than those at 65-days.But,the expression level of longissimus dorsi tissue at the newly born was lower that at the 65-day age.Our results provided the molecular data for the relative study on biological function and genetic evolution in the ADSL gene.