利用CODEHOP设计简并引物克隆红色红曲霉丝氨酸羧肽酶基因片段和序列分析

Degenerate primers design for cloning of serine carboxypeptidase gene fragment of Monascus ruber by CODEHOP software

利用CODEHOP(Consensus-Degenerate Hybrid Oligonucleotide Primers)软件设计了红色红曲霉丝氨酸羧肽酶基因片段的简并引物,选取1对简并引物进行逆转录-聚合酶链式反应(RT-PCR),得到348 bp的聚合酶链式反应产物,经pMD18-T载体克隆转化至大肠杆菌DH5α中,测序后进行BLASTX比对,发现此DNA产物与其他丝氨酸羧肽酶基因序列有相似性,推断所克隆的产物即为红色红曲霉的丝氨酸羧肽酶基因片段.

With the help of CODEHOP, degenerate primers were designed for cloning the serine carboxypepti- dase gene fragment of Monascus tuber. One pair of degenerate primers were selected for RT-PCR, and 348 bp PCR product was obtained, which was transformed into the E. coli DHSot through being linked with pMD18-T vector and sequenced after screening. Similarity alignment showed that the product of the cloned DNA was similar to those of serine carboxypeptidase gene from other strains. The cloned sequence was putatively serine carboxypeptidase gene DNA fragment from M. ruber strain.