Differential Expression Research Of Leizhou Black Duck Pax3, DCN And MSTN Gene In Embryonic Skeletal Muscle Tissue

Duck muscle fiber growth and development is closely related to meat quality traits,Inthe formation and growth of muscle fibers during development,Pax3,DCN and MSTNgene plays a decisive role in physiological characteristics of skeletal muscle.To furtherunderstand the Pax3,DCN and MSTN gene close relationship with the Leizhou black duckin skeletal muscle formation and function during the formation of contact between eachother in skeletal muscle,In order to improve the quality of Leizhou black duck lay themolecular basis.In this study,to observe Leizhou black duck8,13,18,23,28-day-oldembryos chest and leg muscle biopsy and using QRT-PCR and western-blot experiments todetect the developmental expression of Pax3,DCN and MSTN gene in each of the aboveBreaststroke leg in mRNA and protein.At the same time,using QRT-PCR assay a28embryo age Leizhou black duck Pax3,DCN and MSTN gene the difference between theexpression of mind,liver,embryo,lung,muscle stomach,small intestine,brain,breast,leg ofmRNA.In this study,the RT-PCR method for the full length CDS region Leizhou BlackDuck DCN gene sequences were cloned and constructed pcDNA3.1myc-His (-) A-DCNeukaryotic expression vector.The results showed that:（1）From different embryonic age Leizhou black duck breast muscle,leg muscle biopsyshows that the formation of the embryonic skeletal experienced myoblasts, multi-coremuscle control,muscle fiber differentiation process. Leizhou black duck leg embryonicdevelopment significantly earlier than the chest and the Leizhou black duck leg wasprecocious parts for the late part of the pectoral muscle.（2）This study used quantitative PCR method,the differential expression of Pax3genes inembryonic Leizhou black duck skeletal muscle of time and space for testing.The resultsshowed that different embryonic pectoral muscle tissues,Pax3mRNA expression levels ofgenes in the23d reached the peak,when the expression of at least18d; Pax3gene in legmuscle tissue mRNA expression levels peaked in the28d,18d in the expression the leastamount;In the age of28embryos visceral tissues, Pax3gene expression levels in leghighest,followed by the spleen,is again chest.Using western-blot method,the gene expression of Pax3at different embryonic Leizhou black duck pectoral muscle,leg muscletissue levels,and the results showed that the breast muscle tissue,Pax3protein expressionwas highest in the age of23embryos, when the minimum age of13embryos;Leg muscletissue Were highest at age28embryos, the lowest expression of embryonic age8.（3）Differential expression of embryonic DCN genes Leizhou Black Duck time and spacewere detected in skeletal muscle.The results showed that different embryonic pectoralmuscle tissues,DCN mRNA expression levels of genes in the28d reached the peak,whenthe expression of at least8d; DCN in leg muscle tissue mRNA gene expression levelspeaked in the28d,expressed during the8d the least amount;Using western-blot method,thegene expression of DCN in Leizhou black duck different embryonic pectoral and legmuscle tissue levels,and the results showed that,DCN reached the highest expression ofembryonic age of23,in the13embryos the minimum age;Leg muscle tissue Were highestat age28embryos,the lowest expression of embryonic age8.(4) This study used quantitative PCR method, the differential expression of embryonicLeizhou black duck MSTN gene in skeletal muscle were detected in time and space.Theresults showed that different embryonic pectoral muscle tissues,MSTN mRNA expressionlevels of genes in the23d reached the peak,when the expression of at least8d; MSTN genein leg muscle tissue mRNA expression levels peaked in the28d,13d in the expression theleast amount; in visceral tissues of embryonic age28, MSTN gene expression levels in theleg in the highest,followed by the chest.Using western-blot method,the gene expression ofMSTN in Leizhou black duck different embryonic pectoral muscle and leg muscle tissuelevels,and the results showed that, MSTN protein expression reached a maximum at theage of28embryos,embryo age8when reached a minimum;Leg muscle tissue Were highestat age28embryos,the lowest expression of embryonic age8.(5) This study successfully cloned the gene CDS Leizhou Black Duck DCN area andbioinformatics analysis found kinship Leizhou black duck and chicken recent DCN geneand its protein structure prediction,and its protein structure prediction,found Leizhou BlackDuck DCN transmembrane protein does not exist,there is a site of the signal peptide,theprotein is not secreted via the signal peptide is a hydrophobic amino acid sequence with13phosphorylation sites and two N-glycosylation sites.DCN protein contains threeLRR8,1leucine-rich repeat N-terminal domain and a ribonuclease inhibitor belonging tosubfamilies like leucine-rich repeat sequences, involved in the formation of embryonicskeletal muscle regulation; and successfully constructed pcDNA3.1myc-His (-) A-DCNeukaryotic expression vector.