猪白细胞介素-2基因的克隆与表达

Cloning and expression of porcine interleukin-2(IL-2)gene

根据GenBank上发表的猪白细胞介素-2(Interleukin-2,IL-2)基因序列,设计1对引物。采用RT-PCR技术,以ConA刺激的猪外周血淋巴细胞为材料,从总RNA中扩增出484 bp的特异性片段,将其克隆入pGEM-T easy载体。序列测定表明;扩增片段为猪IL-2基因,该基因与GenBank上发表的猪IL-2基因的核苷酸序列同源性达99.8%。将其定向克隆至原核表达载体pET32a,构建了表达重组猪IL-2的基因工程菌株,经IPTG诱导表达和层析纯化,获得了纯化的重组猪IL-2蛋白。

Based on the published nucleotide sequence of porcine interleukin-2 （IL-2）gene, a pair of RT-PCR primers were designed and synthesized. The ConA-stimulated porcine peripheral blood lymphocytes were used as materials,and the specific fragments of 484 bp were amplified from the total RNA and cloned into pGEM-T easy vector by RT-PCR. The sequence determination showed that the homology between the gene of the fragment and the porcine IL-2 gene published in GenBank reached 99.8% in nucleotide sequence. The directional cloning of the fragment into the prokaryotic expression vector pET32a constructed a genetically engineered strain expressing recombinant porcine IL-2 gene, and the purified recombinant porcine IL-2 protein was obtained by induction with IPTG and chromatography.