广西沼泽型水牛白细胞介素-10和干扰素-γ基因的克隆及其序列分析  

Cloning and sequencing of interleukin-10 and interferon-γ gene of Guangxi swamp type buffalo

作　　者：阳玉梅[1] 郑小龙[1] 黄维义[1] 石云良[1] 代华龙[1] 张为宇[1] 

机构地区：[1]广西大学动物科学技术学院,广西南宁530005

出　　处：《黑龙江畜牧兽医》2007年第12期12-16,共5页Heilongjiang Animal Science And veterinary Medicine

基　　金：广西自然科学基金项目(桂科自0007005)

摘　　要：将本地沼泽型成年水牛外周血单核细胞(PBMC)在刀豆素A(ConA)的刺激下体外培养,提取培养的单核细胞总RNA,应用逆转录聚合酶链式反应(RT-PCR)技术扩增水牛白细胞介素-10(IL-10)和干扰素-γ(IFN-γ),并克隆到pMD18-T载体上,用EcoRⅠ和HindⅢ双酶切鉴定筛选阳性克隆,然后进行测序。序列分析结果显示:IL-10开放阅读框(ORF)由536个核苷酸组成,共编码178个氨基酸;在核苷酸水平上与印度水牛、牛、绵羊、猪、马、人和鼠的IL-10基因的同源性为75.9%～99.6%;氨基酸的同源性为74.2%～100%。IFN-γ的ORF由501个核苷酸组成,共编码166个氨基酸;在核苷酸水平上与印度水牛、牛、绵羊、猪、马、人和鼠的IFN-γ基因的同源性为60.7%～99.6%;氨基酸的同源性为40.6%～100%。Abstract： The cDNAs encoding the interleukin- 10 （IL- 10） and interferon- γ（ IFN -γ）of Guangxi swamp type buffalo were amplified by reverse transcription polymerase chain reaction（ RT - PCR） from buffalo peripheral blood mononuclear ceils （PBMC） stimulated concanavalin A（ConA） . The products of RT- PCR were cloned into pMD18 -T vector. Recombinant plasmid were identified by PCR, restriction enzymes digestion and sequencing. The 536 bp of IL - 10 cDNA contain ORF, encoding 178 amino acid. The nucleotide and the deduced amino acide sequences of the buffalo IL - 10（Guangxi） share 75.9% -99.6% and 74.2% - 100% homology with that of buffalo（ India）, bovine,sheep,pig,horse, human and mouse, respectively. IFN - γ contain ORF（501 bp） ,encoding 166 amino acid. The nucleotide and the deduced amino acide sequences of the buffalo IFN- γ（Guangxi） share 60. 7% -99.6% and 40. 6% - 100% homology with that of buffalo（ India）, bovine,sheep,pig ,horse,human and mouse, respectively. It was shown that IL - 10 and IFN - γ of buffalo were successfully. These results will provided foundation for further study on buffalo immunity system and prevention illness using these cytokines.

关 键 词：沼泽型水牛 单核细胞 IL-10 IFN-Γ 克隆 测序 

分 类 号：S823.83[农业科学—畜牧学]