Abstract: The aim of the experiment was to clone the hormone sensitive lipase (HSL) gene of Guangling donkey and analyze its sequence,and further analyze the differential expression level of HSL gene in different tissues of Guangling donkey.In this experiment,RT-PCR method was used to amplify and clone the partial sequence of CDS region of HSL gene of Guangling donkey.After splicing the sequence,the full length sequence of CDS region of HSL gene could be obtained,and a series of bioinformatics analysis was performed on the sequence.Real-time quantitative PCR was used to detect the expression of HSL mRNA in heart,liver,spleen,lung,kidney,longissimus dorsi muscle and subcutaneous fat in Guangling donkey.The results showed that the complete CDS region of HSL gene in Guangling donkey was 2 286 bp in length,encoding a total of 761 amino acids,and the sequence was successfully submitted to NCBI,accession No.MN231003.The homology of the nucleotide sequence of HSL gene in Guangling donkey with the corresponding sequences of Equus caballus,Vicugna pacos,Camelus ferus,Sus scrofa,Bos taurus,Capra hircus,Mus muqulus and Ovis aries were 99.6%,88.9%,88.6%,88.1%,86.9%,85.6%,80.8% and 79.1%,respectively.Phylogenetic tree prediction showed that HSL gene of Guangling donkey was closely related to the horse and the furthest to the mouse.Bioinformatics analysis found that the theoretical isoelectric point of HSL protein was 6.51,and the instability index was 56.83,and the total average hydrophilicity was -0.048,indicating that HSL was an acid-labile water-soluble protein.There were N-terminal domains and α/β hydrolase folding domains as well as regulatory domains in the conserved domains of proteins.There were 88 phosphorylation modification sites and 25 glycosylation modification sites in the protein sequence.There were strong hydrophobic regions in the protein,without signal peptides and transmembrane regions.The secondary structure showed that this protein was composed of alpha-helix,extended chain,beta-turn and random coil,which account for 45.33%,11.70%,5.65%,37.32%,respectively.Real-time quantitative PCR detection results showed that HSL gene mRNA was expressed in 7 kinds of tissues in Guangling donkeys,but there were differences.The highest expression was in subcutaneous fat and the lowest expression in heart,indicating that Guangling donkey HSL gene might be play a very important part in fat deposition in vivo.It provided a theoretical basis for further studying the function of HSL protein and its metabolic regulation mechanism of fat deposition in Guangling donkey.

Key words: Guangling donkey; HSL gene; cloning; bioinformatics analysis; differential expression