猪羰基还原酶1(CBR1)基因克隆及组织表达谱分析

Cloning and expression of porcine carbonyl reductase1(CBR1) gene

为了深入研究猪羰基还原酶1(CBR1)基因的表达调控,通过PCR获得包括猪CBR1基因启动子和外显子、内含子的序列,并进行蛋白结构分析和系统进化关系比较,通过RT-PCR方法检测了CBR1基因在不同组织的表达丰度。结果表明,猪CBR1基因启动子区长度为2 875 bp,具有潜在典型的NFκB等转录因子结合位点;CBR1基因由3个外显子和2个内含子组成,外显子长度分别为289、108和437 bp,其中5′UTR和3′UTR分别为107 bp和242 bp,内含子长度分别为572 bp和3 219 bp,编码区由289个氨基酸组成;该蛋白为亲水性蛋白,无明显的信号肽,有2个跨膜区域;蛋白二级结构和三级结构主要由7个α-螺旋、7个β-折叠以及loop环构成。猪CBR1基因氨基酸序列与人、绵羊氨基酸的相似性较高,为84.48%。q RT-PCR结果表明,怀孕12 d的二花脸子宫内膜中CBR1基因m RNA的表达量极显著高于长大二元母猪;在怀孕12 d母猪9个组织中,CBR1在肾脏中的表达量最高,其次是肝脏和小肠。

The promoter, exons and introns of porcine carbonyl reductasel gene （ CBR1 ） were cloned, and the structure of CBR1 protein was analyzed. The similarity of amino acid was compared with other species and the phylogen of the gene was constructed. The tissues expression of the gene was detected by RT-PCR. The results showed that the promoter of the gene contained 2 875 bp, and typical binding sites of NF K B were predicted. The gene included three exons and two introns, and the formers spanned 289, 108 and 437 bp, respectively. The two introns spanned 572 bp and 3 219 bp, respectively. The 5＇ UTR of 107 bp and 3＇ UTR of 242 bp were identified. The CDS was encoded by 289 amino acids. The CBR1 protein was hydrophilic, and contained two transmemhrane domains but no signal peptide was found. The tertiary structure of the protein was formed by seven α -helixes, seven sheets, and loops. The amino acid sequence of the gene exhibited higher similarity of 84.48% with that of human and sheep. The CBR1 mRNA in the endometrium of Erhualian sows was higher significantly than that of Landrace × large white sows on GD12 （ gestation day 12 ） （ P〈0.01 ） . The gene was expressed with the highest level in kidney tissue and the higher in liver and small bowel tissues of Erhualian on GD12.