| 摘要: |
| 采用同源性克隆和末端快速扩增(RACE)方法, 首次获得全长为1222bp 的条石鲷膜孕激素受体α(mPRα)的cDNA 序列。其开放阅读框为1060bp, 编码了含353 个氨基酸的蛋白, N 端前16 个氨基酸为信号肽。氨基酸序列比较分析发现其存在7 个跨膜区域。其预测的蛋白质三级结构中存在着多个蛋白结合位点。同源性比较和进化分析表明, 条石鲷mPRα 与鲈形目鱼类mPRα 聚为一支, 进化关系较近, 相似度达到95%以上。实时荧光定量PCR 方法检测mPRα mRNA 表达情况: 发现mPRαmRNA 在性成熟雌性条石鲷各组织均有表达, 并在脑、垂体和卵巢组织中的表达较丰富。在条石鲷繁殖周期的脑和垂体组织中, mPRα mRNA的表达水平在卵巢发育的Ⅳ期达到最大值; 在繁殖周期的卵巢组织中, mPRα mRNA 的表达水平从卵巢发育Ⅲ期到Ⅴ期持续升高并且在Ⅴ期达到最大值(P<0.05)。条石鲷雌鱼血清中雌二醇激素的含量变化在整个繁殖周期中差异显著(P<0.05), 在性腺发育Ⅲ期含量迅速升高并在Ⅳ期达到峰值。本研究为条石鲷卵巢成熟调控机制研究提供了基础参考资料。 |
| 关键词: 条石鲷 膜孕激素受体 基因克隆 表达 雌激素 |
| DOI：10.11693/hyhz20140600160 |
| 分类号: |
| 基金项目:国家高技术研究发展计划项目, 2012AA10A413 号; 国家自然科学基金项目, 31201982 号; 国家鲆鲽类产业技术体系,CARS-50 号;山东省优秀中青年科学家科研奖励基金项目, BS2013SW042 号。 |
附件 |
|
| CLONING AND EXPRESSION OF MEMBRANE PROGESTIN RECEPTOR (mPRα) IN ROCK BREAM OPLEGNATHUS FASCIATUS |
|
SHI Bao1, LIU Xue-Zhou1, CHEN Sheng-Yi1,2, XU Yong-Jiang1, ZANG Kun1,2, LI Xiao-Ni1
|
|
1.Key Laboratory of Sustainable Development of Marine Fisheries, Ministry of Agriculture, P. R. China, Qingdao Key Laboratory for Marine Fish Breeding and Biotechnology, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China;2.College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China
|
| Abstract: |
| A full-length cDNA sequence of mPRα was cloned first time from the ovary of Oplegnathus fasciatus by homology cloning and RACE techniques. The mPRα cDNA sequence is 1222bp and its open reading frame is 1060bp, encoding a precursor protein of 353 amino acids preceded by a signal peptide of 16 amino acids residues. Sequence alignment of the mPRα and amino acid of other species suggested that it had seven transmembrance domains. Tertiary structure of mPRα protein showed that it had several binding sites. Sequences comparison and phylogenetic analysis showed the mPRα clusters with perciformes in more than 95% identity. The mPRα mRNA expression of sexually matured rock bream was detected by quantitative real-time PCR (qRT-PCR) method. Tissue expression analysis showed that mPRα mRNA was expressed widely in the fish, and mPRα transcripts were highly abundant in brain, pituitary, and ovary. Expression level of mPRα mRNA in brain and pituitary of reproductive cycle showed that mPRα mRNA were maximum at stageⅤ. And the expression level of mPRα mRNA in the reproductive cycle showed that it significantly increased from Ⅳ ovarian development stage to stageⅤ(P<0.05), then peaked in the stage Ⅴ. Changes in serum estrogen values of reproductive cycle indicated that plasma estrogen values changed dramatically (P<0.05), and the estrogen values increased rapidly and reached maximumat stage Ⅳ. The research may provide a reference for understanding ovarian maturation regulation of O. fasciatus. |
| Key words: Oplegnathus fasciatus membrane progestin receptor gene cloning expression estrogen |
