[Cloning, expression and functional analysis of CuZnSOD gene in swine.]

In order to understand the structure and function of CuZnSOD gene, reveal the effect of the anti-oxidant in swine, and find the molecule marker correlated with meat traits, the cDNA of CuZnSOD gene was cloned and sequenced from muscle of Laiwu black swine by RACE (rapid amplification of cDNA end) techniques. The structure and function of CuZnSOD were analyzed by bioinformatics, and the gene expression profile in different tissues was examined by real-time PCR. The results showed that the full sequence of CuZnSOD cDNA is 658 bp (GU944822), containing 76 bp sequence of 5' UTR and 120 bp sequence of 3' UTR, and coding region (CDS, 462 bp) encodes 153 amino acids. The isoelectric point (pI) of the protein is 6.03, and the molecular weight is 15.9 kDa. There were one O-glycosylation site at the third amino acid and one N-glycosylation site at the eighty-fourth amino acid. The percentage of alpha helix was 1.31%. The alignment similarities of the CDS sequence of swine CuZnSOD with those of cattle, human, rat, and mouse were 87.74%, 87.66%, 83.44%, and 83.23%, and the similarities of amino acid sequence were 90.26%, 94.12%, 92.21%, and 91.50%, respectively. CuZnSOD possesses the typical metal binding ligands (GFHVHQFGDNT). The phylogenic tree based on CuZnSOD protein sequence detected the closest relationship between swine and cattle. CuZnSOD mRNA is a broad-spectrum expression gene, which was detected in brain, heart, spleen, liver, kidney, lung, large intestine, small intestine, spinal cord, muscle, backfat, and stomach. In particular, high expression levels of CuZnSOD mRNA were detected in kidney, small intestine and lung, but low expressions were observed in heart and muscle tissues.