广西巴马小型猪白细胞介素-2基因cDNA的克隆和序列分析

CLONING AND SEQUENCING OF cDNA ENCODING BAMA MINI PORCINE INTERLEUKIN-2 GENE

〔目的〕 对地方猪种广西巴马小型猪淋巴细胞IL - 2基因进行克隆和序列分析 ,为进一步研究猪IL -2基因的功能及其在免疫调节方面的应用奠定基础。〔方法〕 将猪外周淋巴细胞进行体外培养 ,在刀豆球蛋白A(conA)刺激培养 15h后 ,提取培养淋巴细胞总RNA ,应用RT -PCR技术扩增巴马小型猪淋巴细胞白细胞介素 - 2 (IL- 2 )基因 ,并连接到PMD18-T载体上 ,进行测序。〔结果〕 扩增的cDNA全长 5 13个碱基 ,包含 4 6 5个碱基开放阅读框架 ,编码 15 5个氨基酸 ,分子量为 16 .98KD。〔结论〕 此cDNA与人、牛、山羊、绵羊、狗和鼠的IL - 2基因核苷酸比较 ,同源性分别为 85 .2 %、85 .3%、83.4 %、86 .0 %、84 .2 %和 6 9.5 % ,与其他猪种IL - 2基因核苷酸比较同源性为 10 0 %。

The porcine peripheral blood lymphocytes were cultured in vitro. After ConA was used to stimulate the lymphocytes for 15h, total RNA was isolated from the lymphocytes and the mini porcine IL-2 gene was amplified by reverse transcription polymerase chain reaction (RT-PCR). PCR product was cloned into PMD18-T vector and sequenced. Consequently, a DNA clone was obtained encoding Bama mini pig IL-2 of 513bp in length, which contained a complete open reading frame of 456bp. Bama mini Porcine IL-2 composed of 155 amino acids, which a predicted molecular weight of 16977. Bama mini porcine IL-2 has an amino acid homology of 85.2% with human IL-2, 85.3% with bovine , 83.49% with goat, 86% with ovine, 84.2% with dog and 59.5% with rat. And the homology of IL-2 was 100% at nucleotide level between Bama mini porcine and other porcine .