Cloning Of Duck Decorin And Primary Study About Its Function In Muscle Development

Decorin was a member of the small leucine-rich proteoglycan gene family. DCN regulated growth and development of the muscle which went hand in hand with the proteometabolism. Many cell singnaling pathways took part in proteometabolism, such as IGF-1/PI3K/Akt/mTOR and TGFβ. MAFbx, MuRF, mTOR were closely related to muscle proteinmetabolism. Whether there was a relationship between DCN and these genes? There was no report in poultry. DCN was a lingand inhibitor of TGFβ family. DCN played an important role in cell growth, cell differentiation and regulation of connective tissue formation through TGF β, and promoted cell proliferation and differentiation through inhibiting MSTN which was the lingand of TGF β. It was not clear that DCN have same function in poultry. DCN coding domain sequence of duck was obtained from the total RNA of adult duck using RT-PCR method. Then the duck DCN mature peptide cDNA was inserted into pET-32a (+) vector and transformed into Escherichiacoli Rosseta. And then it was induced by IPTG, the recombinant expression products were detected by SDS-PAGE and confirmed by mass spectrum. The relative expression levels of DCN gene in duck different tissues were detected in this research. Meanwhile, in order to further elucidated the role of DCN in duck muscle growth and development process, base on duck muscle cell culture and the DCN gene liposome transfection experiments, this paper discusses the relationships of DCN and ACVR1, ACVR2that are two receports of TGFβ family, DCN and MAFbx, MuRF, mTOR that are closely related to protein metabolism. In order to explain the part function of DCN in duck, provided the theory basis about DCN gene for duck production, and also had some reference for duck breeding and increasing mutton quantity.1) The results of sequence analysis showed that the full duck DCN coding domain sequence was1074bp, encoding for357AA, and the deduced CDS amino acid sequence contained a16AA putative signal peptide.2) The DCN amino acid sequence of duck was composed of nine domains, According to the report about human, there might be certain relationship between LRR4,5and TGFβ.3) Tissue distribution expression of DCN showed different significantly in4weeks ducks. The highest expression was found in lung(p<0.05), followed by small intestine, cardiac, liver, spleen, sebum cutaneum, abdominal fat, then followed by brain, leg muscle, stomach.4) Stability prokaryotic expression system of duck DCN was constructed successfully. The recombinant expression products were purified by Affinity chromatography and detected by SDS-PAGE and confirmed by mass spectroscopy. The molecular weight of DCN protein was54.7kDa.5) Transfer vector of duck DCN gene(pEGFP-N1-DCN) was constructed successfully. Then transfected them into duck myoblasts in vitro. In normal fluorescence logging, transcription level of DCN, mTOR were significantly increased(P<0.05), ACVR1, ACVR2were significantly decreased(P<0.05), but MAFbx, MuRF were not significantly, indicating the role of DCN on regulating the expression of ACVR1, ACVR2and mTOR. DCN might regulate the ACVR1, ACVR2and mTOR.