延边黄牛白细胞介素2的基因克隆和序列分析

Gene cloning and sequence analysis of Interleukin-2 of Yanbian Yellow Calttle

为建立延边黄牛白细胞介素2(IL-2)的基因克隆和序列分析的合理方法体系进行该研究.采集1月龄犊牛静脉血,用密度梯度离心法获取白细胞层,对白细胞进行总RNA的提取,并构建cDNA文库,根据Genbank牛IL-2基因序列(登录号:M12791.1)设计1对特异性引物,用反转录PCR(RT—PCR)技术扩增出目的基因片段,与克隆载体PMD18-T连接,并进行PCR和酶切鉴定,酶切结果为阳性的进行序列分析.结果表明:PCR扩增出大小为261 bp的部分IL-2基因片断,与预期的片段大小一致;此序列与Genbank上牛IL-2从141～401 bp片段的同源性为100%.

Studied the cloning and sequence analysis to build a reasonable method system of the gene. The test retrieval the total RNA from Yanbian calves WBC, reverse transcription to build cDNA library, according to IL-2 gene sequence enunciable in the Gen Bank to design a pair of specificity primer, use PCR technique to amplification purpose fragments, link cloning vectors PMD18-T, PCR and enzyme cuttings appraisement, sequence analysis of the masccline enzyme cuttings. The results showed that the size of PCR amplification IL-2 fragments is 261 bp,conformity for anticipant and coisogenic between cloning IL-2 gene sequences and Gen bank is 100%.