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Cloning, Sequence Analysis, and Prokaryotic Expression of the Porcine DECR1 Gene

摘要2,4-dienoyl-CoA reductase 1(DECR1)is the key rate-limiting enzyme in the metabolism of polyunsaturated fatty acids. Although this protein has been studied in a variety of mammals, its role in por-cine is yet to be fully elucidated. However, it is a candidate determinant/indicator of meat quality,growth traits, and carcass quality. Here, we employed RT-PCR and rapid amplification of cDNA ends (RACE)analysis to amplify the full-length cDNA of DECR1 from Mashen pig liver, and cloned it into the expression vector pET-32a+. After confirmation by sequencing and restriction analysis, the recombinant plasmid was transformed into E. coli BL21 cells. The cDNA of pig DECR1 contained 2,352 nucleotides,including a 987 bp open reading frame flanked by a 53 bp 5'-untranslated region(UTR)and a 1,312 bp 3'-UTR. The pig DECR1 coding sequence encoded 328 amino acid residues, which shared 99%, 88%,87%, 87%, 87%, 87%, and 83% identity with those of Sus scrofa(predicted), Bos taurus, Homo sapiens, Macaca mulatta, Pan troglodytes, Equus caballus, Canis, and Mus musculus, respectively.SDS-PAGE analysis revealed that the recombinant protein was expressed and that the expression level reached its highest level after 4 h induction. Western blot analysis indicated that the molecular weight of the expressed protein was the same as that predicted, ap-proximately 35 kDa. Collectively these data provide the basis for further studies into the physiological functions and molecular mechanisms of the pig DE-CR1 gene.

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分类号 S828
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DOI 10.3969/j.issn.1674-9782.2011.02.061
发布时间 2011-12-15(万方平台首次上网日期,不代表论文的发表时间)
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