China Animal Husbandry and Veterinary Medicine ›› 2022, Vol. 49 ›› Issue (1): 12-22.doi: 10.16431/j.cnki.1671-7236.2022.01.002

• Biotechnology • Previous Articles     Next Articles

Cloning, Sequence Analysis and Tissue Differential Expression of FTO Gene in Guangling Donkey

QIU Lixia1, GUAN Jiawei1, LI Li1, LI Wufeng1, DU Min2   

  1. 1. College of Life Sciences, Shanxi Agricultural University, Taigu 030801, China;
    2. Department of Animal Sciences, Washington State University, Pullman 99164-6310, USA
  • Received:2021-07-27 Online:2022-01-05 Published:2021-12-29

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Abstract: [Objective] The purpose of this study was to clone and analyze the sequence of fat mass and obesity-associated gene(FTO), detect the expression difference of FTO gene in various tissues of Guangling donkey, and explore the effect of the structure of FTO gene related to fat on the physiological and metabolic functions of Guangling donkey. [Method] RT-PCR method was used to amplify and clone the CDS sequence of FTO gene, and analyze the gene and protein function. Meanwhile, Real-time quantitative PCR was used to detect the differential expression of FTO gene in seven tissues (heart, liver, spleen, lung, kidney, longissimus dorsi muscle and subcutaneous fat) of Guangling donkey. [Result] The results showed that the CDS sequence of FTO gene in Guangling donkey was 1 518 bp and could encode 505 amino acids. The sequence was uploaded to NCBI and obtained the accession No.:MZ169553. The similarity of FTO gene in Guangling donkey were 99.3%, 90.3%, 89.5%, 90.8%, 90.7%, 89.2% and 89.2% with Equus caballus, Sus scrofa, Bos taurus, Homo sapients, Vicugna pacos, Ovis aries and Capra hircus, respectively. Phylogenetic tree analysis result showed that FTO gene of Guangling donkey was closely to Equus caballus. The molecular weight of FTO protein was 58.35 ku, the theoretical isoelectric point was 5.07, the fat index was 80.36, the instability coefficient was 48.82, and the average hydrophobic index was -0.550, the FTO protein of Guangling donkey was an unstable acidic hydrophilic protein. There was no signal peptide and transmembrane region of FTO protein, which were mainly located in the cytoplasm, and had 34 phosphorylation sites and 5 glycosylation sites. The secondary structure prediction of FTO protein showed that α-helix (43.96%) and random coil (37.82%) were the main structures. Real-time quantitative PCR analysis showed that FTO gene was all expressed in 7 kinds of tissues, the expression of FTO gene in lung and subcutaneous fat were extremely significantly higher than other tissues (P<0.01), and the expression was the lowest in longissimus dorsi muscle. [Conclusion] The results of this study provided a solid theoretical basis for the further research on gene expression, mechanism of fat deposition, and improved the meat quality of Guangling donkey.

Key words: Guangling donkey; FTO gene; cloning; bioinformatics analysis; tissue expression

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