企鹅珍珠贝Sox9基因的克隆及表达分析

Molecular Cloning and Expression Analysis of Sox9 Gene from Pteria penguin

【目的】更好地了解企鹅珍珠贝性别转换机制。【方法】RACE-PCR技术克隆企鹅珍珠贝Sox9基因cDNA的全长序列,分析其理化性质和进化地位;荧光定量PCR技术分析Sox9基因在各组织及不同时期性腺中的表达特征。【结果与结论】Sox9基因cDNA序列全长2 267 bp,其中开放阅读框(ORF)为1 410 bp,编码469个氨基酸。氨基酸序列比对显示企鹅珍珠贝Sox9基因与黑蝶真珠蛤(Pinctada margaritifera)和马氏珠母贝(Pinctada fucata)有高度同源性(>81%)。Sox9在企鹅珍珠贝各组织中均有表达,在足中表达量最高(P<0.05),精巢中其次;Sox9在成熟期精巢检测到最大表达量(P<0.05),在发育早期精巢、退化期精巢和成熟期卵巢中表达量较低,其中发育早期卵巢表达量最低(P<0.05)。

【Objective】To better understand gender conversion mechanism of Petria penguin.【Method】Full-length cDNA of Sox9 gene from Pteria penguin was cloned by RACE-PCR and its physicochemical properties and evolutionary status were analyzed. Fluorescent quantitative PCR was used to analyze the expression of Sox9 gene indifferent tissues and gonads at different life cycle stages.【Result and conclusion】The full-length cDNA of Sox9 gene was 2 267 bp, with the longest open reading frame（ORF） of 1 410 bp, It encoded for a protein of 469 amino acids. Amino acid sequence alignment showed that the Sox9 gene in Pteria penguin shared highly sequence identity with Sox9 geneof Pinctada margaritifera and Pinctada fucata（ 81%）. The realtime-PCR results showed that Sox9 was expressed in all tissues. The expression level was highest in foot（P〈0.05）, followed by testis. The expression analysis in gonads of different periods indicated that Sox9 had the highest expression in mature testes, but lower expression in early testis, emission testis and mature ovary, with the lowest level in early ovary.