昆明鼠白细胞介素-15基因的克隆与序列分析

Cloning and Sequence Analysis of IL-15 Gene of Chinese KM Mice

参照GenBank登录的小家鼠(Mus musculus)白细胞介素-15基因序列设计引物,利用RT-PCR技术对刚地弓形虫RH株诱导昆明(KM)鼠72 h后的脾细胞进行扩增,克隆得到了KM鼠的成熟IL-15核苷酸序列。经测序分析发现,KM鼠成熟IL-15序列长度为345 bp,编码114个氨基酸,与小家鼠IL-15序列相似性达到100%。序列比对和遗传进化分析表明,KM鼠成熟IL-15序列与挪威鼠亲缘关系较近,核苷酸和氨基酸序列相似性分别为94.2%和95.5%;与土拨鼠、人、灵长类、兔、犬、猫、牛、野猪等哺乳动物亲缘关系相对较远,核苷酸和氨基酸序列相似性分别为71.2%～78.8%和69.5%～74.7%;与珍珠鸟、非洲蟾蜍和斑马鱼亲缘关系最远,核苷酸和氨基酸序列相似性分别为6.1%～50.9%和16.1%～32.5%。KM鼠IL-15基因的成功克隆为进一步研究其功能及免疫增强作用奠定了基础。

According to IL-15 gene sequence of Mus musculus available in GenBank database,specific primers were designed and synthesized.Using reverse transcription-polymerase chain reaction（RT-PCR） method,mature IL-15 sequence was amplified from the spleen cells of Kunming（KM） mice stimulated for 72 h by Toxoplasma gondii RH strain,then cloned into pMD18-T vector and transformed into the competent Escherichia coli JM109 for obtaining the positive clone and sequenced.Sequencing analysis revealed that the length of mature IL-15 of KM mice was 345 bp,encoding 114 amino acids.The results of sequence alignment and genetic evolution analysis showed that the KM mice mature IL-15 sequence was the closest to Norvegicus rattus,and the homologies of nucleotide and amino acid sequence were 94.2% and 95.5% respectively.Sequence similarity between the KM mice mature IL-15 sequence and that of Cavia porcellus,human,primates,Oryctolagus cuniculus,dog,cat,Bos taurus,Sus scrofa etc.,and the nucleotide homology ranged between 71.2% to 78.8%,and the amino acid homology was 69.5% to 74.7%.The successful cloning of KM mouse IL-15 gene provided foundation for further studies of its functions and immune stimulating effects.