巴马香猪CIDEa基因克隆及组织表达分析

doi:10.16431/j.cnki.1671-7236.2021.12.002

Abstract

Abstract: The purpose of this experiment was to clone the CIDEa gene sequence of Bama Xiang pig, predict its structure and function, and analyze its tissue expression.According to the CIDEa gene sequence of Sus scrofa published by NCBI (accession No.: NM_001112696.2), primers were designed with Oligo 7.0 software, and the CIDEa gene sequence of Bama Xiang pig was amplified and cloned by RT-PCR.The nucleotide sequence and the hydrophobicity, physicochemical properties, transmembrane domain, secondary structure, tertiary structure and interaction proteins of the encoded proteins were analyzed by bioinformatics software.The expression of CIDEa gene in subcutaneous fat, liver, kidney, lung, spleen, longissimus dorsi muscle and heart of Bama Xiang pigs were detected by Real-time quantitative PCR.The results showed that the CDS regions of CIDEa gene in Bama Xiang pig was successfully obtained, which was 660 bp in length, encoded 219 amino acid.The nucleotide similarity with Sus scrofa, Bos taurus, Equus caballus, Canis lupus familiaris, Homo sapiens, Macaca mulatta and Mus musculus in GenBank were 99.4%, 84.7%, 81.5%, 80.9%, 79.7%, 78.9%, and 76.1%, respectively.Compared with the nucleotide sequence of Sus scrofa CIDEa in GenBank, there were four base mutations, of which A609G and T627C were synonymous mutations, and C173T (Pro→Leu) and C631T (Arg→Cys) were missense mutations.The molecular weight of CIDEa protein in Bama Xiang pig was 24 483.57 u, the isoelectric point was 9.48, and the instability index was 52.86, indicating that the protein was unstable.The protein contained the highest Leu (13.2%) and the lowest Trp (0.5%).CIDEa protein in Bama Xiang pig was an extramembrane hydrophilic protein, which contains a superfamily domain (CIDE-N).The secondary structure prediction of the protein showed that the percentage of alpha helix, beta turn, extended chain, random coil of CIDEa protein in Bama Xiang pig was 45.66%, 5.94%, 14.61% and 33.79%, respectively.The predicted results of the tertiary structure were consistent with those of the secondary structure.Protein interaction analysis showed that CIDEA protein of Bama Xiang pig interacted with PPARG, PPARGC-1, COX8H, PRDM16, DIO2, TMEM26, ELOVL3, COX7A1, CIDEC, DFFB proteins.The results of Real-time quantitative PCR showed that the expression of the CIDEa gene was the highest in subcutaneous fat of Bama Xiang pig and significantly higher than that in other tissues (P<0.05), and the lowest expression in heart.The results provided a theoretical basis for further exploring the regulatory mechanism of CIDEa gene on fat deposition in Bama Xiang pig.

Key words: Bama Xiang pig; CIDEa gene; cloning; expression

CLC Number:

S828.8⁺9

LUO Yunyan, WEI Chongwan, LIU Chunyan, LIU Gangyi, JIANG Qinyang, HUANG Yanna. Cloning and Tissue Expression of CIDEa Gene in Bama Xiang Pig[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(12): 4339-4347.

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Cloning and Tissue Expression of CIDEa Gene in Bama Xiang Pig

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