盘羊与巴什拜羊杂交后代SPLUNC1基因克隆及序列分析

Cloning and sequence analysis of SPLUNC1 gene from hybrid of Argali and Bashibai Sheep

为了克隆盘羊与巴什拜羊杂交后代短鄂、肺及鼻咽上皮细胞克隆1(SPLUNC1)基因cDNA全长序列,本研究根据GenBank上已公布的牛、山羊的SPLUNC1基因序列来设计简并引物,以盘羊与巴什拜羊杂交F1代为材料,克隆出盘羊与巴什拜羊杂交后代SPLUNC1基因的片段序列,再利用RACE法克隆其SPLUNC1基因3′端和5′端,测序后拼接获得SPLUNC1基因全长cDNA序列。结果显示,盘羊与巴什拜羊杂交后代SPLUNC1基因的cDNA全长为1117 bp,5′非编码区(UTR)为84 bp,3′UTR为265 bp,开放阅读框(ORF)为768 bp,编码255个氨基酸。预测该基因编码蛋白的等电点5.006,分子量26.57 kDa。系统进化分析表明,盘羊与巴什拜羊杂交后代SPLUNC1氨基酸序列与绵羊的氨基酸序列同源性最高。本研究克隆了盘羊与巴什拜羊杂交后代SPLUNC1基因全长cDNA序列,为后续深入研究该杂交后代SPLUNC1基因的功能奠定基础。

This paper aimed to clone Argali hybrid offspring short palate,lung and nasal epithelium clone 1(SPLUNCl) gene,and obtain the cDNA full-length sequence.The sequences of cow and goat SPLUNCl gene from the GenBank were used to design the degenerate primers,and mucosa of oral palate were scraped from Argali hybrid offspring F1.Rapid Amplification cDNA ends(RACE) technology was used to clone Argali hybrid offspring SPLUNCl gene fragment sequence.The results showed that the cDNA length of Argali hybrid offspring SPLUNCl gene was 1117 bp,including 84 bp in 5' region(UTR) and 265 bp in 3' regions(UTR).The Open Reading Frame was 768 bp encoding 255 amino acids.It was predicted that the protein encoding by the gene with a calculated molecular weight of 26.57 kD and an isoelectric point of 5.006.Phylogenetic analysis indicated that Argali hybrid offspring shared the high homology with Ovis aries.Argali hybrid offspring SPLUNCl gene was cloned in this study,which laid a foundation for the further studies on the function of SPLUNCl gene in Argali hybrid offspring.