籽鹅卵巢产蛋性能相关基因全长cDNA序列的克隆与分析

目的克隆并分析籽鹅卵巢产蛋性能相关基因EST1的全长cDNA序列。方法利用实时荧光定量PCR技术对EST1基因在籽鹅产蛋前期与产蛋期卵巢中mRNA表达水平进行检测,并采用RACE(Rapid amplification of cDNAends,RACE)技术对该基因全长cDNA序列进行克隆,应用生物信息学预测方法对其编码的蛋白质进行分析。结果籽鹅产蛋期卵巢组织中EST1基因mRNA的表达水平显著高于产蛋前期(P<0.05)。经RACE技术获得EST1基因全长cDNA序列长1715bp,具有单一的完整开放阅读框(ORF,14～1318bp),推测编码蛋白含434个氨基酸残基,相对分子质量为107100,等电点为5.00。该蛋白为细胞质内蛋白,含3个跨膜螺旋,蛋白序列中含1个信号肽切割位点。结论经分子生物学软件进行蛋白质功能预测,初步确定EST1基因为籽鹅α-烯醇化酶蛋白基因,推测该基因可能参与籽鹅产蛋性能的分子调控。

Cloning and Analysis of Full-length cDNA Sequence of A Gene Associated with Egg Production in Ovary Tissue of Laying Goose

Objective To clone and analyze the full-length cDNA sequence of EST1 gene associated with egg production in ovary tissue of laying goose.Methods The expression levels of EST1 mRNA in the ovary tissue of laying goose in pre-laying and laying periods were determined by real-time fluorescent quantitative PCR,based on which the full-length cDNA sequence of EST1 gene was cloned by RACE(rapid amplification of cDNA ends)technique,and the encoded protein was analyzed by bioinformatic predictive method.Results The expression level of EST1 mRNA in the ovary tissue of laying goose in laying period was significantly higher than that in pre-laying period(P < 0.05).The full-length cDNA sequence of EST1 gene obtained by RACE technique,at a length of 1 715 bp,contained a single open reading frame(ORF,14 ～ 1 318 bp).The encoded protein deduced consisted of 434 amino acid residues,with a relative molecular mass of 107 100 and an isoelectric point of 5.00.The protein located in cytoplasm contained three transmembrane coiling,of which the sequence contained a signal peptide cleavage site.Conclusions EST1 gene was preliminarily identified as α-enolase protein gene by the prediction of function with molecular biological software,and was deduced to be involved in the molecular regulation of egg production of laying goose.