应用RACE技术扩增驴DGAT1基因3′端及序列分析

摘要

DGAT1（acyl CoA： diacylgycerol acyltransferase） gene is the most important functional candidate gene for milk production traits.Because the bovine DGAT1 gene has 98%homology with the predicted Equus animals.In this experiment,based on bovine DGAT1 gene,we used the RNA of donkey lacteal gland tissue to make the template and designed the special primers according to different species＇ DGAT1 homology.The experiment obtained the amplified fragment using PCR and RACE technique.The amplified fragment was purified and inserted into pUCmT Vector then transformed into colibacillus cells.The positive recombinant plasmid was extracted and was sequenced to analyse its sequence.The result showed that this sequence was consistent with expected aim,we have firstly cloned the 3＇ end cDNA sequence of donkey DGAT1 gene by the homology analysis of other animals.%DGAT1（脂酰辅酶A：二脂酰甘油酰基转移酶）基因是产奶性状的一个重要功能侯选基因。由于通过预测奶牛DGAT1基因序列与马属动物DGAT1基因序列具有98%的同源性,本试验从影响牛产奶性状的DGAT1基因出发,以驴乳腺组织的RNA为模板,按照不同物种DGAT1基因的相似性设计特异引物,运用PCR和3′RACE技术扩增并获得了特异片断,特异片段回收纯化连接到pUCmT Vector载体后,转化到大肠杆菌中并筛选阳性菌落;提取质粒进行测序,结果发现该段序列与预期的目标一致,通过与其他动物同源性比较分析说明已首次克隆到驴DGAT1基因3′端。