嗜酸乳杆菌亚油酸异构酶基因的克隆与表达

Cloning and Expression of Conjugated Linoleic Acid Isomerase Gene of Lactobacillus acidophilus

利用分子生物学方法对嗜酸乳杆菌AS1.1854菌株的亚油酸异构酶基因进行克隆与表达.培养菌体后提取总DNA,用PCR法扩增其亚油酸异构酶基因,再将其克隆到pET30a载体上,转入到大肠杆菌BL21株中表达,在低温下诱导表达出重组蛋白,并用Ni2+金属鳌合层析对重组蛋白进行分离和纯化.通过实验,得到了亚油酸异构酶的基因,成功转入到载体中,表达出了可溶性的重组蛋白,并对其进行了纯化.实验表明,在原核细胞中可以表达出可溶性重组亚油酸异构酶,以200mmol/L的咪唑洗脱液进行洗脱可以获得目标蛋白.

This paper deals with the cloning and expression of conjugated linoleic acid isomerase gene of Lactobacillus acidophilus AS1. 1854 by means of molecular biological method. In the investigation, first, the genome was extracted after the cell was cultured and gathered, and was amplified by means of PCR. Next, the amplified gene was cloned into pET30a plasmid vector and subsequently converted into Escherichia coli BL21 strain. Then, the gene was induced to. express the recombinant protein at a low temperature, and the recombinant protein was separated and purified with Ni-NTA agarose. Finally, the target gene was amplified and converted into plasmid, and the recombinant protein was expressed and separated through the experiment. It is found that soluble recombinant conjugated linoleic acid isomerase can be expressed in prokaryotic cell, and that the target protein can be obtained by washing with 200 mmol/L imidazole.