山羊ACSS2基因的克隆、序列分析及组织表达研究

doi:10.16431/j.cnki.1671-7236.2020.02.003

摘要/Abstract

摘要： 本研究旨在对山羊乙酰辅酶A合成酶2（acetyl-CoA synthetase 2，ACSS2）基因进行克隆和生物信息学分析，并检测其在山羊不同泌乳时期乳腺组织中的表达量变化。以山羊乳腺组织RNA为模板，采用RT-PCR方法扩增并克隆山羊ACSS2基因完整CDS区序列，对测序结果进行生物信息学分析，并对ACSS2基因在山羊不同泌乳时期乳腺组织中的表达量进行分析。结果显示，山羊ACSS2基因CDS区序列长2 106 bp，编码701个氨基酸；山羊ACSS2基因与牛、马、人、犬、猪、小鼠和鸡的同源性分别为97.8%、92.0%、91.3%、91.3%、91.1%、88.1%和73.3%。蛋白理化性质分析结果表明，ACSS2蛋白分子质量为78.72 ku，理论等电点为6.03，属于酸性蛋白；跨膜结构和信号肽分析表明，ACSS2蛋白不含跨膜结构和信号肽；结构域分析表明，该蛋白含有1个乙酰辅酶A合成酶N端结构域。亚细胞定位分析结果表明，该蛋白主要分布在内质网（44.4%）、线粒体（33.3%）、细胞质（11.1%）和细胞核（11.1%）中。蛋白质结构预测发现ACSS2蛋白含有α-螺旋（29.10%）、延伸链（21.54%）、β-转角（9.84%）及无规则卷曲（39.52%）。实时荧光定量PCR分析结果表明，ACSS2基因在不同泌乳时期均有表达，其中在泌乳中期表达量最高，在干奶期表达量最低。本试验结果为进一步研究山羊ACSS2基因在脂质代谢过程中的功能及转录调控机制提供了参考。

关键词: 山羊; ACSS2基因; 克隆,生物信息学分析; 表达分析

Abstract: This study was aimed to clone acetyl-CoA synthetase 2(ACSS2) gene in goats,perform bioinformatics analysis,and detect its mRNA expression level in mammary gland during different lactation stages in goats.Mammary gland tissue RNA in goats was used as a template,and the complete CDS region of ACSS2 gene in goats was amplified and cloned.The sequencing result was analyzed by bioinformatics,and the expression of ACSS2 gene in mammary gland tissues during different lactation stages in goats were analyzed.The results showed that the total CDS region sequence of ACSS2 gene in goats was 2 106 bp,which encoded 701 amino acids.The homology of ACSS2 gene in goats with Bos taurus,Equus caballus,Homo sapiens,Canis lupus familiaris,Sus scrofa,Rattus norvegicus and Gallus gallus were 97.8%,92.0%,91.3%,91.3%,91.1%,88.1% and 73.3%,respectively.The physicochemical properties of ACSS2 protein showed that its molecular weight was 78.72 ku,and its theoretical isoelectric point was 6.03,which belonged to an acidic protein.No transmembrane structure and signal peptide was found in ACSS2 protein,but it contained a N-terminal domain of acetyl-CoA synthetase.Subcellular localization analysis indicated that ACSS2 protein was located in endoplasmic reticulum (44.4%),mitochondria (33.3%),cytoplasm (11.1%) and nucleus (11.1%).ACSS2 protein structure was mainly consisted of alpha helix (29.10%),extended chain (21.54%),beta turn(9.84%) and random coil (39.52%).Real-time PCR results revealed that the highest and the lowest expression of ACSS2 gene in goats were in the middle lactation and dry lactation periods,respectively.These results provided references for further study of the function and transcriptional regulation mechanism of ACSS2 gene in lipid metabolism of goats.

Key words: goat; ACSS2 gene; cloning; bioinformatics analysis; expression analysis

中图分类号:

宋国华, 颜铎, 李瑞婷, 许会芬, 李明. 山羊ACSS2基因的克隆、序列分析及组织表达研究[J]. 中国畜牧兽医, 2020, 47(2): 336-344.

SONG Guohua, YAN Duo, LI Ruiting, XU Huifen, LI Ming. A Cloning,Sequence Analysis and Tissue Expression of ACSS2 Gene in Goats[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(2): 336-344.

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