| 曼氏无针乌贼精子鞭毛蛋白1(Spef1)基因克隆以及组织表达特异性 |
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| 引用本文： | 周林,李颖,吕振明,迟长凤,吴常文,史会来.曼氏无针乌贼精子鞭毛蛋白1(Spef1)基因克隆以及组织表达特异性[J].水产学报,2018,42(8):1189-1198. |
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| 作者姓名： | 周林 李颖 吕振明 迟长凤 吴常文 史会来 |
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| 作者单位： | 浙江海洋大学海洋科学与技术学院国家海洋设施养殖工程技术研究中心海洋生物种质发掘与利用国家地方联合实验室;浙江省海洋水产研究所 |
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| 基金项目： | 浙江省自然科学基金（LY15C190010）；舟山市科技专项（2016C41016）；浙江省科研院所专项（2015F50055）；浙江海洋大学“海洋科学”省重中之重学科开放课题（20160116） |
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| 摘 要： | 为研究精子鞭毛蛋白1(Spef1)在精子鞭毛结构的形成与组装中的生物学意义及功能,本研究采用RACE技术和荧光定量PCR技术对曼氏无针乌贼Spef1(简称Sj Spef1)基因cD NA全长进行克隆和组织表达特异性分析。结果显示,SjS pef1 cD NA全长序列共1 135 bp,5′和3′非编码区分别为178 bp和165 bp,预测的开放阅读框(ORF)全长792 bp。编码的蛋白理论分子量为30.567 7 ku,等电点7.03,是一种亲水性蛋白。不存在跨膜区以及信号肽序列,是在细胞内发挥作用的蛋白。二级结构分析发现该蛋白含有丰富的螺旋结构(49%)。氨基酸同源建模显示其蛋白的CH2结构域主要由4个螺旋结构组成,并由多个loop结构串联而成。同源氨基酸序列比对发现,它与加州双斑蛸的相似性最高且仅为59.49%,表明Spef1在进化中并不保守。基于Spef1氨基酸序列构建的系统进化分析表明,曼氏无针乌贼和加州双斑蛸进化关系最近。组织特异性分析表明Spef1在曼氏无针乌贼的精巢中有显著表达。Spef1基因的成功克隆以及组织表达特异性分析对于深入研究其细胞定位以及生物学功能具有重要意义。
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| 关 键 词： | 曼氏无针乌贼 Spef1 cDNA 生物信息学 荧光定量PCR |
| 收稿时间： | 2017/4/10 0:00:00 |
| 修稿时间： | 2017/11/1 0:00:00 |
Sperm flagellar protein 1 (Spef1) gene clone and expression analysis in Sepiella japonica |
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| ZHOU Lin,LI Ying,L&#; Zhenming,CHI Changfeng,WU Changwen and SHI Huilai.Sperm flagellar protein 1 (Spef1) gene clone and expression analysis in Sepiella japonica[J].Journal of Fisheries of China,2018,42(8):1189-1198. |
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| Authors: | ZHOU Lin LI Ying L&#; Zhenming CHI Changfeng WU Changwen and SHI Huilai |
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| Institution: | National and Provincial Joint Laboratory of Exploration and Utilization of Marine Aquatic Genetic Resources, National Engineering Research Center of Marine Facilities Aquaculture, School of Marine Science and Technology, Zhejiang Ocean University, Zhoushan 316022, China,National and Provincial Joint Laboratory of Exploration and Utilization of Marine Aquatic Genetic Resources, National Engineering Research Center of Marine Facilities Aquaculture, School of Marine Science and Technology, Zhejiang Ocean University, Zhoushan 316022, China,National and Provincial Joint Laboratory of Exploration and Utilization of Marine Aquatic Genetic Resources, National Engineering Research Center of Marine Facilities Aquaculture, School of Marine Science and Technology, Zhejiang Ocean University, Zhoushan 316022, China,National and Provincial Joint Laboratory of Exploration and Utilization of Marine Aquatic Genetic Resources, National Engineering Research Center of Marine Facilities Aquaculture, School of Marine Science and Technology, Zhejiang Ocean University, Zhoushan 316022, China,National and Provincial Joint Laboratory of Exploration and Utilization of Marine Aquatic Genetic Resources, National Engineering Research Center of Marine Facilities Aquaculture, School of Marine Science and Technology, Zhejiang Ocean University, Zhoushan 316022, China and Marine Fisheries Research Institute of Zhejiang, Zhoushan 316021, China |
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| Abstract: | To study the biological significance and function of sperm flagellar protein 1 (Spef1) in the formation and assembly of spermatozoa, rapid amplification of cDNA ends (RACE) and qRT-PCR techniques were used to clone the gene cDNA of the Spef1 in Sepiella japonica and analyse the tissue expression specificity. The results showed that a 1 135 bp full-length cDNA of Spef1 gene from S. japonica (described as SjSpef1) was obtained, which consisted of a 178 bp 5'' untranslated region (UTR) and a 165 bp 3''UTR. The predicted open reading frame (ORF) was 792 bp, molecular weight of the deduced protein was 30.567 7 ku, and its pI was 7.03. The SjSpef1 was hydrophilic, without signal peptide sequence and transmembrane regions, and it might play a intracellular role. Secondary structure analysis revealed that it contained rich spiral structures (49%). Homology modeling indicated that CH2 domain of Spef1 mainly consisted of four spiral structures, which were connected by some loop structures. The multiple sequence alignment showed that SjSpef1 had the highest similarity with Octopus bimaculoides which was 59.49%, which showed that Spef1 was not very conserved in evolution. Phylogenetics analysis based on Spef1 demonstrated that S. japonica had the closest relationship with O. bimaculoides. SjSpef1 gene was expressed significantly in testis by qRT-PCR. Its cloning and analysis of tissue specific expression will be of great significance to exploring its cellular localization and biological function in the future. |
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| Keywords: | Sepiella japonica Spef1 cDNA bioinformatics qRT-PCR |
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