摘要
从水貂脑垂体中分离提取总RNA,经过RT-PCR分别扩增出水貂生长激素前体肽和成熟肽cDNA。PCR产物经凝胶回收纯化,克隆到载体pGEM-T,然后转化感受态细胞DH5α。在含X-galI、PTG的LB(Amp+)平板上筛选阳性克隆,提取质粒作限制性酶切鉴定后,对目的片段进行测序。结果表明,RT-PCR扩增出的2个cDNA片段碱基数分别为713bp和780bp,用DNAsis2.5软件进行分析表明,此扩增序列与GenBank中发表的水貂生长激素cDNA100%相同。前体肽编码区由651bp组成,编码216个氨基酸,包括长度为29个氨基酸的信号肽;成熟肽的开放阅读框全长564bp,编码187个氨基酸,分子量约为21kDa。通过Blast比对,分析了与人、马、牛、猪、山羊、绵羊、猫、狗、大鼠和小鼠生长激素核苷酸序列的同源性。
According to the published cDNA sequence of Mink Growth Hormone (mGH) in GenBank, two pairs of primer mGH were designed with DNAsis2.5 software. From the total RNA of mink pituitary, cDNA sequences of mGH precursor and mature peptide were obtained by RT-PCR. The obtained cDNA sequences were respectively cloned to pGEM-T vector. The sequencing results showed that length of mGH precursor cDNA fragment was 651bp. ORF included of 564bp, encoding 187 amino acids, and molecular weight was about 21kDa. Compared the cDNA of mGH mature peptide with those of human, horse, bovine, pig, goat, ovine, cat, dog, rat and mouse, the identities respectively were 73%?93%?88%?98%?88%? 88%?95%?95%? 89% and 90%.
出处
《特产研究》
2005年第2期1-4,共4页
Special Wild Economic Animal and Plant Research
关键词
水貂
生长激素CDNA
克隆
序列分析
<Keyword>mink
growth hormone cDNA
cloning
sequence analysis
