大熊猫核糖体蛋白L10亚基基因的cDNA克隆及序列分析  

cDNA cloning and sequence analysis of ribosomal protein L10 gene in giant panda

作　　者：苏秀兰[1] 侯怡铃[1] 李俊[1] 孙冰[1] 吴光富[1] 宋嫣[1] 张田[1] 侯万儒[1] 

机构地区：[1]西华师范大学生命科学学院,四川南充637002

出　　处：《第三军医大学学报》2010年第15期1617-1620,共4页Journal of Third Military Medical University

基　　金：国家自然科学基金(30470261);四川省应用技术项目(2006J13-057);四川省教育厅重点科研项项目(07ZA120);四川省教育厅青年基金(09ZB088);四川省重点学科动物学建设资助项目(404001);四川省科技厅应用基础项目(2009JY0061)~~

摘　　要：目的了解大熊猫(ailuropoda melanoleuca)核糖体蛋白L10亚基基因(RPL10)的结构特点及其与已报道的人和其他哺乳动物核糖体蛋白亚基RPL10基因的异同。方法根据已报道的部分哺乳动物RPL10基因的相关信息设计引物,运用RT-PCR技术,从大熊猫的肌肉组织总RNA中对核糖体蛋白亚基RPL10基因的进行克隆、测序;采用Genescan软件,对所克隆的基因序列进行氨基酸序列推定;采用ORF finder软件进行DNA序列的开放阅读框查找;采用DNAMANVersion 6软件,对基因序列和氨基酸序列进行同源性比较;采用Ex2PASy Proteomics Server软件,进行蛋白质功能位点和生化特性预测分析;采用http://www.imtech.res.in/raghava/apssp2/对蛋白质二级结构进行模拟分析。结果大熊猫RPL10基因的表达序列长为685 bp,开放阅读框(ORF)为645 bp,编码214个氨基酸的蛋白质,该蛋白的相对分子质量为24.599 7×103,等电点为10.74,含有4个功能位点,分别为:2个蛋白激酶C磷酸化位点,2个N化位点,2个酰胺化位点,1个核糖体蛋白L10e信号位点。进一步分析发现,该基因的表达序列及其编码的氨基酸序列与已报道的人、牛、家犬、褐家鼠、小家鼠有很高的相似性,其表达序列同源性分别为92.4%、92.4%、97.21%、88.84%与89.61%;其编码的氨基酸序列除与牛的同源性为99.07%外,与其他哺乳动物同源性为100%。而且,其蛋白质的高级结构除牛外其他物种也具有一致性。结论无论是从RPL10基因的序列、结构、功能位点,还是从其编码的氨基酸的序列和二级结构进行分析,都表明大熊猫的RPL10基因和已报道的哺乳动物的RPL10基因具有高度的遗传稳定性和功能一致性。Objective To study the structure of ribosomal protein L10 （RPL10） gene in giant panda （ailuropoda melanoleuca） and its similarities and difference with the structures of other reported human and mammalian RPLIO genes. Methods Primers were designed according to the reported data on the structure of mammalian subunit RPLIO gene. RPLIO gene was cloned from the total RNA in muscle tissue of giant panda using RT-PCR technique and then sequenced with the GenScan software. The sequences of amino acid and cloned gene were compared using DNAMAN Version 6. The functional sites and biochemical properties of RPLIO were analyzed using the Ex2PASy Proteomics Server. The secondary structure of RPLIO was simulated and analyzed as described at the Website （http ：//www. imteeh, res. in/raghava/apssp2）. Results The length of RPLIO gene in giant panda was 685 bp containing an ORF of 645 bp, encoding 214 amino acids. The relative molecular weight of RPLIO was 24. 599 7 × 10^3 with a PI of 10.74, 4 functional sites （2 protein kinase C phosphorylation sites, 2 N-myristoylation sites, 2 amidation sites, and 1 RPL10e signature site）. Further analysis showed that the expression rate of homologous sequences in RPL10 of humans, cattle, dogs, brown rabbits and small rabbits was 92.4% , 92.4% , 97.21% , 88.84% and 89.61% , respectively, which were highly homologous to those （ 100% and 99.07% ） in cattle and other mammals. Moreover, the structure ef RPL10 was also similar in all species but not in cattle. Conclusion The sequence, structure, and functional site of RPL10 gene or itencoded amino acid sequence and secondary structure analysis indicate that the genetic stability and function of RPL10 gene in giant panda are consistent with those of RPL10 gene reported in mammals.

关 键 词：大熊猫 RT-PCR RPL10 克隆 序列分析 

分 类 号：Q343.35[生物学—遗传学] Q75