| 绵羊GlyCAM-1基因克隆、核苷酸变异及其组织表达分析 |
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| 引用本文： | 郝志云,王继卿,胡江,刘秀,李少斌,罗玉柱. 绵羊GlyCAM-1基因克隆、核苷酸变异及其组织表达分析[J]. 核农学报, 2020, 34(8): 1690-1697. DOI: 10.11869/j.issn.100-8551.2020.08.1690 |
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| 作者姓名： | 郝志云 王继卿 胡江 刘秀 李少斌 罗玉柱 |
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| 作者单位： | 甘肃农业大学动物科学技术学院/甘肃省草食动物生物技术重点实验室/甘肃省牛羊基因改良工程实验室,甘肃 兰州 730070 |
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| 基金项目： | 安徽师范大学培育项目;国家自然科学基金;甘肃省基础研究创新群体项目 |
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| 摘 要： | 糖基化依赖细胞粘附分子(GlyCAM-1)是粘液素(mucin)糖蛋白家族的成员,是乳腺细胞合成的一种乳脂球膜蛋白(MFGMPs)的重要组成部分。GlyCAM-1在乳腺发育和泌乳中发挥着重要作用,为探究该基因的序列特征、核苷酸序列变异和表达特征,以高泌乳量的小尾寒羊(泌乳高峰期和空怀期)和低泌乳量的甘肃高山细毛羊(泌乳高峰期)的乳腺组织为研究对象,利用克隆测序、RT-qPCR、生物信息学等方法克隆绵羊GlyCAM-1基因的编码区,分析GlyCAM-1的理化性质和蛋白质结构,并检测GlyCAM-1基因的组织表达特性。结果表明,绵羊的GlyCAM-1基因CDS区全长465 bp,编码154个氨基酸。测序结果表明,在该基因CDS区检测到7个SNPs,其中2个为同义突变,5个为错义突变。GlyCAM-1二级结构主要以α-螺旋和无规则卷曲为主,延伸链则散布于整个蛋白质结构中。String分析结果表明,GlyCAM-1蛋白与CD34、MadCAM-1都作为L-选择素(L-selectin)的配体发挥作用;RT-qPCR结果表明,GlyCAM-1基因表达具有组织特异性、品种特异性和时空特异性,乳腺、心脏、肝脏、肺脏、脾脏、肾脏、卵巢和背最长肌8个组织中,GlyCAM-1基因只在乳腺和肺脏组织中表达,在其余6个组织中均不表达,其中乳腺中的表达量最高;在泌乳高峰期的乳腺组织中,GlyCAM-1在高泌乳量的小尾寒羊中的表达量显著高于甘肃高山细毛羊(P<0.05);在小尾寒羊的乳腺组织中,GlyCAM-1在泌乳高峰期的表达量极显著高于空怀期(P<0.01)。本研究结果丰富了绵羊基因组数据,为深入研究GlyCAM-1基因的泌乳生物学功能及其机理提供了基础数据。
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| 关 键 词： | GlyCAM-1基因 乳腺 绵羊 克隆 组织表达 SNPs |
| 收稿时间： | 2018-12-18 |
Gene Cloning,Nucleotide Variation and Tissue Expression Analysis of GlyCAM-1 in Sheep (Ovis aries) |
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| HAO Zhiyun,WANG Jiqing,HU Jiang,LIU Xiu,LI Shaobin,LUO Yuzhu. Gene Cloning,Nucleotide Variation and Tissue Expression Analysis of GlyCAM-1 in Sheep (Ovis aries)[J]. Acta Agriculturae Nucleatae Sinica, 2020, 34(8): 1690-1697. DOI: 10.11869/j.issn.100-8551.2020.08.1690 |
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| Authors: | HAO Zhiyun WANG Jiqing HU Jiang LIU Xiu LI Shaobin LUO Yuzhu |
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| Affiliation: | College of Animal Science and Technology/Gansu Key Laboratory of Herbivorous Animal Biotechnology/Gansu Engineering Lab of Genetic Improvement in Ruminants, Gansu Agricultural University, Lanzhou, Gansu 730070 |
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| Abstract: | The glycosylation-dependent cell adhesion molecule (GlyCAM-1) is a member of the mucin glycoprotein family and is an important component of milk fat globule membrane protein (MFGMPs) synthesized by mammary epithelial cells. GlyCAM-1 plays an important role in mammary gland development and lactation. To explore the sequence characteristics, nucleotide sequence variation and expression characteristics of GlyCAM-1 gene, the mammary gland tissue of high-lactating small-tailed Han sheep (lacking peak and empty period) and low lactation Gansu alpine fine-wool sheep (the peak of lactation) were used to study. The coding region of sheep GlyCAM-1 gene was cloned by cloning sequencing. The physicochemical properties and structure of GlyCAM-1 protein were analyzed by bioinformatics and the tissue expression characteristics of the GlyCAM-1 gene were detected by RT-qPCR. The results showed that the CDS of GlyCAM-1 gene in sheep was 465 bp in length and encoded 154 amino acids. The sequencing results showed that 7 SNPs were identified in the CDS region of the gene, of which 2 were synonymous mutations and 5 were missense mutations. The secondary structure of GlyCAM-1 is predominantly composed of α-helix and random coil, and the extended chain is dispersed throughout the protein structure. Results of String analysis indicated that GlyCAM-1 protein interacts with CD34 and MadCAM-1 as ligands of L-selectin. RT-qPCR results indicate that the expression of GlyCAM-1 gene is tissue-specific, breed-specific and space-time specific. Among the eight tissues of the mammary gland, heart, liver, lung, spleen, kidney, ovary and longissimus dorsi. GlyCAM-1 gene was only expressed in mammary gland and lung tissues, but not in the other six tissues. and the expression level in the mammary gland is higher. In the mammary gland at the peak of lactation, the expression of GlyCAM-1 level in the high-lactating small-tailed Han sheep was significantly higher than that in the Gansu alpine fine-wool sheep (P<0.05). In the mammary gland of Small Tail Han sheep, the expression at the peak of lactation was significantly higher than that in the empty period (P<0.01). The results of this study enriched the sheep genome data and provided basic data for further study on the lactation biological function and mechanism of GlyCAM-1 gene. |
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| Keywords: | GlyCAM-1 gene mammary gland sheep clone tissue expression SNPs |
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