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角蛋白18(K18)是角蛋白的一种,为中间纤维蛋白,是细胞骨架的必要组成部分。采用RACE法克隆了大鳞副泥鳅K18基因cD NA的全长序列,并运用实时荧光定量PCR技术探讨其在不同组织中的表达。结果表明,克隆得到的大鳞副泥鳅K18基因cD NA序列全长为1 718 bp,其中包含1个长1 296 bp开放阅读框,编码431个氨基酸,蛋白质相对分子量48.66 ku。同源性分析显示,大鳞副泥鳅K18与泥鳅的相似性最高为98%,与斑马鱼、金鱼、虹鳟、白斑狗鱼等物种K18也存在不同程度的相似性。系统进化分析表明,大鳞副泥鳅与其他物种的K18系统发育同源,并与泥鳅的关系最为密切。通过实时荧光定量PCR分析,K18基因在大鳞副泥鳅的肠、肌肉、胃、心脏、肝脏、精巢、卵巢、脾脏等8种组织中都有表达,其中,肠表达最高,心脏最低。研究旨在为大鳞副泥鳅的分子生物学研究提供参考资料,并为更好地了解大鳞副泥鳅细胞骨架结构系统提供依据。
Keratin 18 (K18), a type of keratin, is an intermediate fibrin that is an integral part of the cytoskeleton. The full-length cDNA of cD NA of K18 gene was cloned by RACE, and its expression in different tissues was explored by real-time fluorescence quantitative PCR. The results showed that the cD NA sequence of K18 gene from Paralichthys leyi showed a total length of 1 718 bp, including a 1 296 bp open reading frame encoding 431 amino acids with a relative molecular mass of 48.66 ku. Homology analysis showed that the similarity of Paralichthys olivaceus K18 to loach was 98%, which was similar to K18 of zebra fish, goldfish, rainbow trout and white spotted fish. Phylogenetic analysis showed that Paramisgurnus dabryanus shared the same K18 phylogeny with other species and had the closest relationship with Loach. Real-time quantitative PCR analysis showed that K18 gene was expressed in eight kinds of tissues, including gut, muscle, stomach, heart, liver, testis, ovary and spleen of Paramisgurnus dabryanus, with the highest expression in the intestine and the lowest in the heart. The aim of this study is to provide references for the molecular biology of Paramisgurnus dabryanus and to provide a basis for better understanding the cytoskeleton structure of Paramisgurnus dabryanus.