[1]张欣如,古丽米热·阿不都热依木,陈莹,等.绵羊基质Gla蛋白基因克隆、表达谱及其在卵巢组织的定位[J].江苏农业学报,2024,(07):1276-1284.[doi:doi:10.3969/j.issn.1000-4440.2024.07.014]
　ZHANG Xinru,Gulimire·Abudureyimu,CHEN Ying,et al.Cloning, expression profile and ovarian localization of sheep matrix Gla protein gene[J].,2024,(07):1276-1284.[doi:doi:10.3969/j.issn.1000-4440.2024.07.014]
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绵羊基质Gla蛋白基因克隆、表达谱及其在卵巢组织的定位()
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江苏农业学报[ISSN:1006-6977/CN:61-1281/TN]

卷:
期数:
2024年07期
页码:
1276-1284
栏目:
畜牧兽医·水产养殖·益虫饲养
出版日期:
2024-07-30

文章信息/Info

Title:
Cloning, expression profile and ovarian localization of sheep matrix Gla protein gene
作者:
张欣如1; 古丽米热·阿不都热依木1;  陈莹1; 马秀玲2; 林嘉鹏1; 汪立芹1; 黄俊成1; 吴阳升1
(1.农业农村部草食家畜遗传育种与繁殖重点实验室/新疆动物生物技术重点实验室/新疆畜牧科学院生物技术研究所,新疆乌鲁木齐830011;2.新疆农业大学动物科学学院,新疆乌鲁木齐830052)
Author(s):
ZHANG Xinru1; Gulimire·Abudureyimu1; CHEN Ying1; MA Xiuling2; LIN Jiapeng1; WANG Liqin1; HUANG Juncheng1; WU Yangsheng1
(1.Key Laboratory of Genetics Breeding and Reproduction of Grass Feeding Livestock, Ministry of Agriculture and Rural Affairs/Key Laboratory of Animal Biotechnology of Xinjiang/ Institute of Biotechnology, Xinjiang Academy of Animal Science, Urumqi 830011, China;2.College of Animal Science, Xinjiang Agriculture University, Urumqi 830052, China)
关键词:
基质Gla蛋白; 绵羊; 卵巢; 颗粒细胞; 黄体
Keywords:
matrix Gla protein; sheep; ovary; granular cell; corpus luteum
分类号:
S826.2
DOI:
doi:10.3969/j.issn.1000-4440.2024.07.014
文献标志码:
A
摘要:
本研究旨在克隆绵羊基质Gla蛋白(MGP)基因的蛋白质编码序列(CDS),制备特异抗体,并检测该基因在绵羊卵巢组织中的表达分布。以卵泡总cDNA为模板,扩增获得绵羊MGP基因CDS区全长序列;合成绵羊MGP蛋白C末端15个氨基酸长度的多肽,免疫小鼠,制备抗血清;利用实时荧光定量PCR(qPCR)和免疫印迹技术分析其在不同组织和不同大小的卵泡中的表达水平;通过免疫组织化学方法分析其在卵巢组织中的表达分布特征。结果表明,成功获得绵羊MGP基因的CDS区全长片段(467 bp),编码103个氨基酸,理论相对分子量为12 230.96,等电点为9.27;N末端19个氨基酸序列被预测为信号肽序列,成熟肽有84个氨基酸，为外泌性蛋白质,无胞内区; MGP总蛋白质氨基酸序列与山羊、牛的同源蛋白质相似性分别为100.0%、99.0%,与人、小鼠的相似性均为85.4%。MGP mRNA和蛋白质在绵羊心、肝、脾、肺、肾等组织中的相对表达量较低,在卵巢、输卵管、子宫等生殖系统或组织中的相对表达量较高,在黄体中的相对表达量也较高;MGP在卵巢组织颗粒细胞、卵丘细胞中均有表达,但在闭锁卵泡中不表达。本研究结果可为进一步研究MGP在卵巢组织中的生物学功能提供参考。
Abstract:
The aim of this study was to clone the protein coding sequence (CDS) of sheep matrix Gla protein (MGP) gene to prepare specific antibody and to detect the expression distribution of the gene in sheep ovarian tissues. Total sheep follicle cDNA was used as the template to amplify and obtain the full-length sequence of the CDS region of sheep MGP gene. Peptide with a length of 15 amino acids from the C-terminal end of the sheep MGP protein was synthesized and was used to immunize mouse and perpare antiserum. Expression levels of MGP in different tissues and follicles of varying sizes were analyzed by real-time fluorescence quantitative PCR (qPCR) and immunoblotting technology. Immunohistochemistry method was used to analyze the expression and distribution characteristics of MGP in ovarian tissue. The results revealed that the overall fragment (467 bp) of sheep MGP gene CDS region was obtained successfully, and the fragment encoded 103 amino acids, with a theoretical relative molecular weight of 12 230.96 and an isoelectric point of 9.27. Sequence composed of 19 amino acids from the N-terminal was predicted as the signal peptide sequence, and the mature peptide contained 84 amino acids, which was extra-membranous domain protein without intracellular region. The total protein amino acid sequences of MGP shared similarities with those of the homologous proteins from goats and cows of 100.0% and 99.0% respectively, and the similarity with human and mouse was 85.4%. The relative expression levels of MGP mRNA and protein were low in sheep tissues such as heart, liver, spleen, lung and kidney, but were high in reproductive systems or tissues, such as ovary, fallopian tube and uterus, and the relative expression level was also high in corpus luteum. The expression levels of MGP granulosa cells and cumulus cells were high, whereas it didn’t expressed in atresia follicles. The study result can provide a basis for further investigation of the biological role of MGP in ovarian tissues.

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备注/Memo

备注/Memo:
收稿日期:2023-08-25基金项目:新疆维吾尔自治区自然科学基金项目(2021D01A59)作者简介:张欣如(1997-),女,新疆哈巴河人,硕士,助理研究员，研究方向为动物遗传育种与繁殖。(E-mail)1635907063@qq.com通讯作者:吴阳升,(E-mail)victorwys@hotmail.com
更新日期/Last Update: 2024-09-14