绵羊SUN2基因生物信息学分析及其在A549细胞中的表达

doi:10.16431/j.cnki.1671-7236.2021.11.003

Abstract

Abstract: This study was aimed to clone Sad1 and UNC84 domain containing 2(SUN2) gene in sheep and conduct bioinformatics analysis, construct eukaryotic expression vector and detect its expression in A549 cells. The specific primers were designed according to sequence of SUN2 gene in sheep (GenBank accession No. :XM_015095026.2), and the CDS sequence of SUN2 gene in sheep was amplified by RT-PCR method and cloned. After sequencing and identification, the sequence were analyzed by bioinformatics, and pEGFP-C1-SUN2 recombinant plasmid was constructed. The pEGFP-C1-SUN2 recombinant plasmid was transfected into A549 cells by liposome transfection method and its expression was detected. The results showed that the total length of SUN2 gene CDS was 2 187 bp, encoding 728 amino acids, the theoretical molecular mass was 81.06 ku, the molecular formula was C₃₅₉₃H₅₆₃₉N₁₀₃₁O₁₁₁₀S₁₄, the isoelectric point was 6.20, the total number of atoms was 11 387, and the instability coefficient was 56.88, which was unstable protein. The similarity of the nucleotide sequence of SUN2 gene between sheep and Capra hircus, Bostaurus, Susscrofa, Equuscaballus, Canis lupus familiaris, Rattus norvegicus, Mus musculus and Homo sapiens were 98.9%, 97.2%, 91.6%, 90.6%, 87.1%, 82.4%, 82.1% and 86.1%, respectively. The gene similarity among different species was relatively high and conservative in the evolution process. The phylogenetic tree results showed that sheep were closely related to Capra hircus, Bos taurus, Sus scrofa, Equus caballus and Canis lupus familiaris, but far related to Gallus gallus. The prediction of structural domain and transmembrane domain showed that SUN2 protein in sheep contained a highly conserved SUN domain, and there was a transmembrane domain. The signal peptide prediction showed that there was no signal peptide site, and the hydrophobicity analysis was a hydrophilic protein. The secondary structure of the amino acid sequence was mainly alpha helix (51.65%), and the rest were random coil (31.46%), extended chain (12.36%) and beta turn (4.53%). The eukaryotic expression vector pEGFP-C1-SUN2 of SUN2 gene in sheep was successfully constructed, which was transfected into A549 cells and the protein expression was detected. The results would provide a reference for the functional study of SUN2 gene in sheep, and laid a foundation for the subsequent exploration of the role of SUN2 gene in ovine pulmonary denocarcinoma.

Key words: sheep; SUN2 gene; vector construction; bioinformatics analysis; transfection

CLC Number:

DU Xiaoyue, ZHANG Liang, LI Huiping, LIU Shuying. Bioinformatics Analysis of SUN2 Gene in Sheep and Its Expression in A549 Cells[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(11): 3929-3939.

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Bioinformatics Analysis of SUN2 Gene in Sheep and Its Expression in A549 Cells

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