草原红牛Acot2基因克隆及其在各组织中的表达差异研究

doi:10.16431/j.cnki.1671-7236.2019.11.003

Abstract

Abstract: This experiment was aimed to study the gene function of Acot2 in Red steppe using bioinformatics,and investigate the mRNA expression of Acot2 gene in different tissues of Red steppe.Primers were designed according to the sequence of cattle Acot2 gene (accession No.:NM_001101938.1) in GenBank.The complete CDS region of Acot2 gene was cloned by RT-PCR and sequencing.Sequence homology alignment and phylogenetic tree of Acot2 gene constructed by software.The corresponding amino acid sequence was obtained.The physical and chemical property,subcellular structure,hydrophilicity and hydrophobicity and phosphorylation site were analyzed by online prediction software.Protein secondary and tertiary structures were predicted by DNAStar software and SWISS-MODEL.Real-time PCR was used to detect the mRNA expression of Acot2 gene in different tissues of Red steppe.The results showed that the length of Acot2 gene CDS was 1 395 bp,encoding 464 amino acids.Acot2 gene CDS in Red steppe had higher homology with Bubalus bubalis(98.3%),but lower homology with Macaca mulatta and Pan troglodytes (80.5% and 80.4%).The molecular formula of Acot2 protein was C₂₃₁₇H₃₆₀₆N₆₄₀O₆₂₈S₁₄,the molecular weight was 50.924 ku,the theoretical isoelectric point was 8.84.Most of the amino acid residues were hydrophilic residues,and the total average hydrophilicity was -0.094.The results of subcellular localization showed that Acot2 protein was located in the endoplasmic (30.4%),mitochondria (26.1%),golgi (17.4%),cytoplasmic (17.4%),vacuolar (4.3%) and nuclear (4.3%).There were 20 phosphorylation sites.The main forms of the secondary structure were alpha helix (21.8%),beta turn (33.4%),beta fold (18.4%) and random coil (26.4%),which was the same as the prediction of the tertiary structure.Real-time PCR result showed that the expression of Acot2 gene in Red steppe was the highest in stomach,it was rarely expressed in lung.The Acot2 gene was lowly conserved in the process of biological evolution.Acot2 protein was structurally stable,and it belonged to water soluble protein.The protein played a role in mitochondria and endoplasmic.The expression of Acot2 gene in different tissues of Red steppe was significant difference.This study provided the basis for further investigation of the effects of Acot2 gene on lipid metabolism in livestock and the screening of quality candidate genes in Red steppe.

Key words: Red steppe; Acot2 gene; cloning; bioinformatics analysis; expression difference

CLC Number:

LIU Lixiang, GAO Yi, LYU Yang, XUE Jiajia, HU Zhongchang, ZHANG Guoliang. Cloning of Acot2 Gene and Its Expression Difference of Different Tissues in Red Steppe[J]. China Animal Husbandry and Veterinary Medicine, 2019, 46(11): 3154-3162.

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Cloning of Acot2 Gene and Its Expression Difference of Different Tissues in Red Steppe

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