西农关中奶山羊PDCD4基因的克隆与组织表达分析

Cloning and tissue expression analysis of PDCD4 in Guanzhong dairy goat

旨在研究程序性细胞死亡因子4(PDCD4)基因与奶山羊乳腺发育的关系。应用RT-PCR技术克隆PDCD4基因编码区(CDS)序列,并进行生物信息学分析,运用荧光定量PCR(qRT-PCR)技术检测不同组织mRNA相对表达量。结果表明:克隆获得的关中奶山羊PDCD4基因CDS序列长1 410 bp,编码469个氨基酸,分子质量为51.8 ku,分子式为C_(2245)H_(3606)N_(656)O_(230)S_(79),理论等电点为5.03;编码蛋白含有4个N-糖基化位点、6个O-糖基化位点及53个潜在的磷酸化位点;PDCD4蛋白为亲水性酸性蛋白质,无信号肽,无跨膜结构域;系统进化树显示PDCD4蛋白的氨基酸序列在不同物种间相似度较高,奶山羊与绵羊、牛、猫、猪、弯角大羚羊、美洲白尾鹿等同源性均在90%以上,其中与绵羊的亲缘关系最近;蛋白互作分析显示PDCD4蛋白能与EIF4A1、EIF4G1、PTEN、EIF4A2、EIF4B、RECK、GMPS、EEF1G等蛋白相互作用;qRT-PCR结果显示PDCD4基因在奶山羊各组织中均有表达,在乳腺组织中的表达量最高,其次为肺,在肌肉组织中几乎不表达。本研究为山羊乳腺功能的研究提供了一定的参考资料。

This experiment was to study the relationship between the PDCD4 gene and dairy goats mammary gland development. The sequence of the CDS region of the PDCD4 gene was cloned using the RT-PCR technology, and bioinformatics analysis was performed. The relative expression of mRNA in different tissues was detected using qRT-PCR. The CDS region of PDCD4 obtained by cloning was 1 410 bp, encoding 469 amino acids. Its molecular weight was 51.8 ku, its molecular formula was C_(2245)H_(3606)N_(656)O_(230)S_(79), and its theoretical isoelectric point was 5.03. The encoding protein contained 4 N-glycosylation sites, 6 O-glycosylation sites and 53 potential phosphorylation sites. PDCD4 was a hydrophilic acidic protein with no signal peptide or transmembrane domain. Its phylogenetic tree showed that the amino acid sequence of the PDCD4 protein was similar among different species. Its homology between dairy goat and sheep, cattle, cats, pigs, elantelope, American white-tailed deer and so on was over 90%, and the closest genetic relationship with sheep was found. The protein interaction analysis showed that PDCD4 could also interact with proteins such as EIF4 A1, EIF4 G1, PTEN, EIF4 A2, EIF4 B, RECK, GMPS, and EEF1 G. The qRT-PCR results showed that PDCD4 was expressed in different tissues of dairy goats, and its highest expression in the mammary gland, followed by the lung, but it hardly expressed in the muscle tissue. The present study provided a reference for research on the function of the mammary gland of goats.