牙鲆dazl基因的克隆与表达分析

发布时间：2018-01-26 00:35

本文关键词： 牙鲆 dazl cDNA克隆基因表达　出处：《海洋渔业》2016年04期 　论文类型：期刊论文

【摘要】：dazl(deleted in azoospermia-like)基因是多种物种精子发生的重要调控因子,为探讨该基因在牙鲆(Paralichthys olivaceus)性腺发育分化中的作用,本研究采用同源克隆和RACE技术克隆了牙鲆dazl基因的cDNA序列,利用生物信息学方法分析了其序列结构,并利用实时荧光定量PCR技术检测了该基因在牙鲆成体组织和早期发育阶段的表达情况。结果发现:牙鲆dazl cDNA序列全长2 031 bp,包括105 bp的5'端非翻译区,1 275 bp的3'端非翻译区和编码217个氨基酸残基的651 bp开放阅读框;氨基酸同源序列比对显示该序列存在一个高度保守的RRM(RNA recognition motif)结构域,且该结构域在鱼类中具有100%的序列一致性;荧光定量PCR结果显示dazl基因主要在牙鲆性腺中表达,且在精巢中的表达高于卵巢;在牙鲆早期发育阶段,dazl基因在未受精卵、受精卵及囊胚期均有较高的转录本,而在原肠胚期表达开始降低,至孵化后3 d一直保持较低的水平,这些丰富表达的母源性dazl mRNA可能参与了胚胎发育中原始生殖细胞的形成。本研究为进一步阐明dazl基因在牙鲆生殖发育中的功能奠定了基础。
[Abstract]:Dazl(deleted in azoospermia-like gene is an important regulator of spermatogenesis in many species. To investigate the role of this gene in the development and differentiation of gonad of Paralichthys olivaceus in Paralichthys olivaceus. In this study, the cDNA sequence of dazl gene of Paralichthys olivaceus was cloned by homologous cloning and RACE technique, and its sequence structure was analyzed by bioinformatics. The expression of the gene in the adult tissues and early development stage of Paralichthys olivaceus was detected by real-time fluorescence quantitative PCR. The results showed that the dazl cDNA sequence of Paralichthys olivaceus was 2031 BP in length. It includes a 5'untranslated region of 105 BP and a 3'terminal untranslated region of 1 275 BP and an open reading frame of 651 BP encoding 217 amino acid residues. The amino acid sequence alignment showed that the sequence had a highly conserved RRM(RNA recognition motif domain. The domain has a sequence consistency of 100% in fish. Fluorescence quantitative PCR showed that dazl gene was mainly expressed in the gonads of Paralichthys olivaceus and was higher in the testis than in the ovaries. In the early stage of development of Paralichthys olivaceus, there were high transcripts of dazl gene in unfertilized eggs, fertilized eggs and blastocysts, but the expression of dazl gene began to decrease at the stage of proembryo, and remained at a low level until 3 days after hatching. These abundant expression of maternal dazl. MRNA may be involved in the formation of primordial germ cells in embryonic development. This study lays a foundation for further elucidating the function of dazl gene in the reproductive development of Paralichthys olivaceus (Paralichthys olivaceus).
【作者单位】：上海海洋大学水产与生命学院农业部淡水水产种质资源重点实验室;
【基金】：国家自然科学基金青年项目(41306128);国家自然科学基金面上项目(30271017)
【分类号】：S917.4
【正文快照】： DAZ基因家族包括daz、dazl(daz-like)和bolue基因,这3者的编码蛋白均为RNA结合蛋白,在生殖细胞中特异表达,参与调节生殖细胞的发育和分化[1-2],是精子发生过程中减数分裂的主要调控因子[3]。bolue和dazl是常染色体基因,一般认为bolue是DAZ基因家族的祖先基因(ancestral gene),

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