猪脂蛋白脂酶基因片段的克隆及不同体重的表达差异  

Cloning of Lipoprotein Lipase(LPL)Gene of Swine and the Difference of LPL Gene Expression at Different Avoirdupois Stages

作　　者：单体中[1] 汪以真[1] 李民[1] 

机构地区：[1]浙江大学饲料科学研究所动物分子营养学教育部重点实验室,杭州310029

出　　处：《农业生物技术学报》2006年第2期151-155,共5页Journal of Agricultural Biotechnology

基　　金：国家重点基础研究发展计划(973)项目(No.2004CB117506)资助

摘　　要：从杜长大母猪的肠系膜脂肪中提取基因组RNA,用RT-PCR扩增脂蛋白脂酶(lipoproteinlipaseLPL)基因,获得1条约689bp的片段,以pGEM-TEasyvector为载体,将该基因片断克隆到大肠杆菌(Escherichiacoli)DH5α中。从筛选的阳性克隆中分离出LPL基因,测定其序列。分析表明,该片段为LPLcDNA的部分序列,编码229个氨基酸组成的多肽。研究得到的基因片段与报道的猪脂肪组织中LPLcDNA部分序列同源性达到98%,氨基酸序列同源性达到99.1%。以LPL基因片段的克隆为基础,构建了优化的半定量RT-PCR法,以β-actin为内标,研究不同体重猪脂肪组织LPL基因表达的差异。结果发现,从刚出生到30kg,LPL基因表达呈上升趋势,在30～50kg,LPL基因表达呈下降趋势,50～90kg,LPL基因表达又呈上升趋势。Genome RNA was extracted from mesentery adipose of pig （Duroc Landrace Yorkshire） and iipoprotein iipase （LPL） mRNA was amplified using RT-PCR. A DNA fragment about 689 bp in length was obtained and the PCR product was cloned into pGEM-T vector. The LPL gene was isolated and sequenced from the screened positive clones. Result of sequence analysis showed that this fragment was the partial sequence of LPL eDNA and coded 279 amino acid residues. The gene homology of the obtained fragment compared with that of reported LPL gene sequence in adipose of porcine was up to 98%, and amino acid homology was 98.94%. Based on the LPL gene clone, an optimal semi-quantitative RT-PCR method was successfully constructed. Using β-actin as inner controi, the difference of LPL gene expression at different avoirdupois of swine was researched, which increased from new born to 30 kg, decreased from 30 to 50 kg and rose again at the stage of 50 to 90 kg.

关 键 词：猪 脂蛋白脂酶(LPL)基因 肠系膜脂肪 RT-PCR 

分 类 号：S185[农业科学—农业基础科学]