牙鲆emx1基因的克隆及表达

doi:10.13560/j.cnki.biotech.bull.1985.2016.06.023

摘要/Abstract

摘要： 为阐述空通气孔同源框1(empty spiracles homeobox 1, emx1)基因在牙鲆发育中的作用，通过PCR技术克隆牙鲆emx1基因，运用BioEdit、MEGA等软件分析其序列结构特征，并采用荧光定量PCR技术分析其在牙鲆早期发育和组织分布中的表达情况。结果成功获得一长度为1 127 bp的牙鲆emx1 cDNA序列，其中包括708 bp的开放阅读框，编码235个氨基酸。同源性比对发现，牙鲆emx1基因的氨基酸序列与雀鲷和花鳉同源性最高，为95%，与其它物种如斑马鱼、墨西哥脂鲤、原鸡、热带爪蟾、小鼠和人的同源性分别为87%、84%、83%、80%、72%和72%。系统进化树分析显示，牙鲆Emx1蛋白与鱼类Emx1紧密聚为一支。荧光定量 PCR显示，emx1基因在牙鲆卵巢中的表达量丰富，精巢中次之，而在其他各组织的表达量则较低。emx1在从原肠胚到孵化后10 d的牙鲆早期发育中也发现了相对较高的表达，且在胚孔封闭和心跳期有最高的表达。结果表明emx1基因很可能在牙鲆早期发育和生殖系统中发挥重要作用。

关键词: 空通气孔同源框1(emx1), 牙鲆, cDNA克隆, 基因表达

Abstract: To explore the function of the empty spiracles homeobox 1(emx1)gene in the development of Paralichthys olivaceus，the emx1 was cloned by PCR，the sequence feature was analyzed by BioEdit and MEGA software，and the expression in early development and adult tissues was detected by real-time quantitative PCR. The obtained emx1 cDNA was 1 127 bp，including a 708 bp open reading frame(ORF)，encoding a polypeptide of 235 amino acids. The alignment analysis of amino acid sequence discovered that the emx1 from P. olivaceus showed the highest 95% homology with that of Stegastes partitus and Poecilia formosa，the sequence similarities between P. olivaceus and Danio rerio, Astyanax mexicanus, Xenopus tropicalis, Gallus gallus, Mus musculus and Homo sapiens，were 87%，84%，83%，80%，72% and 72%，respectively. The Emx1 protein was highly homologous with that in other fish through phylogenic analysis. Real-time quantitative PCR revealed that the emx1 mRNA was expressed in all detected tissues of P. olivaceus. The expression of emx1 was the highest in the ovary and secondly highest in the testis，while relatively low in other tissues. In early development stage，i.e.，from gastrula to 10 d after hatching，a relatively high expression of the emx1 mRNA was also observed，and the highest expression occurred in the blastopore stage and heart-beating stage. The findings indicated that the emx1 played an important role in early development and gonad development stages of P. olivaceus.

Key words: emx1, Paralichthys olivaceus, cDNA cloning, gene expression

孙文慧, 张俊玲, 施志仪, 孙近近, 向玉婷, 陈晓武. 牙鲆emx1基因的克隆及表达[J]. 生物技术通报, 2016, 32(6): 161-167.

SUN Wen-hui, ZHANG Jun-ling, SHI Zhi-yi, SUN Jin-jin, XIANG Yu-ting, CHEN Xiao-wu. Identification and Expression Analysis of emx1 from Paralichthys olivaceus[J]. Biotechnology Bulletin, 2016, 32(6): 161-167.

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