The MRNA Expression Of Four Candidate Genes For Muscle Development In Chicken With The Differentiation Of Myoblasts And Development Of Muscles

Meat production rate of chickens is an economically important trait. Filtering,authentication related functional genes are fundamental to molecular assisted breeding. Inour previous study，the results of genome-wide association studies（GWAS）on the carcasstraits using Illumina60k SNP chip revealed that some SNPs and candidate genes weresignificantly (or suggestive linkage) associated with breast and leg muscles’ weight ofBeijing-You chicken. In this study, the variation of mRNA expression of4candidate genesin muscle tissues including the proliferation and differentiation of myoblasts and also indifferent stages of growth was identified. To clarify its relationship with muscle growth,this study aims to identify major genes for these traits and may lead to the development oftechniques for marker assisted selection.1. The analysis on the tissue-specificity of the genes. The variations of mRNAexpression of4candidate genes in10tissues including myocardium, breast muscle et al. inBeijing-You chicken were compared with each other s. Results showed that the expressionof NCOA7, MYCN and HDAC in the leg muscle was significantly higher than other tissues（P<0.05）.GJA1also indicated a highexpression in gizzard and heart tissues.2. The temporal and spatial expression of characteristics of the late embryonic periodto98th days after hatch. We choose BJY chicken as experiment ma terials. At each of4stages of embryonic development (e14, e16, e18, and e21d), the breast and leg musclesfrom a group of6BJY chickens were collected. And from hatch to98th days (0,28,56,70,84,98days), the breast and leg muscles from a group of10BJY chickens at each periodwere collected. mRNA expression were detected using Q-PCR. Results showed that themRNA expression levels of NCOA7in both breast and thigh muscles were higher in e18dduring e14d to e21d and started the expression decline after e18d. NCOA7mRNAexpression in breast muscle showed after rising has a downward trend during hatch to98thdays, and56D showed a highest expression level. The mRNA expression levels of MYCNwere continued to rise during e14d to e21d. The expression in breast a nd thigh muscles atdevelopment stages from hatch to98th day were reduced markedly (p<0.01) on0-28daysof age where after28D, the expressions were not significant, and remained at relativelylow levels. HDAC2had a similar trend with MYCN. The GJA1gene’s mRNA expressionlevels was raising during embryonic development stages. There were no significantdifferences during hatch to28D, but after28D, GJA1mRNA expression levels were continued to rise.3. The study on the proliferation and differentiation properties of primary chickenembryonic myoblasts cultured in vitro.16days SPF chick embryos were used to isolatechicken embryonic myoblasts and culture until72h. Collection of cells and detection ofindividual gene’s expression were performed at proliferative stage. After cells’ grow to80%, DMEM medium was replaced with5%horse serum to induce differentiation andcollected the myoblast cells from12to120hours, and then detect genes expressions. Theresults showed that the expression of NCOA7, MYCN and HDAC2genes in chickenembryonic myoblasts at proliferative stage was significantly lower than the expression ofcells at late differentiate phase (P<0.05). As the extensionof time, the3genes’expressionswere changed and shown highest at96h and started to decline after that.GJA1geneexpression at proliferative stage was significantly higher than differentiate stage(P<0.01)and peak expression was observed at96h.4. Valproic acid (VPA) inhibited the expression of HDAC2gene on proliferation anddifferentiation of myoblast in-vitro. Normal myoblast cells in culture are divided into twogroups, one was control and another one VPA added with the concentration of1mol/mL.The regulation of HDAC2and specific muscle growth factors was detected, such as PAX7and other genes expressions during the proliferative and differentiation. The resultsshowed that，after adding VPA， the PAX7gene was up regulated. The cell proliferativeability of VPA group was significantly higher than control group during0-72h. Theexpression of HDAC2in the group which added VPA was lower than in the control groupduring each of the differentiation times (12,24,48,72,96and120h). The expression ofMYOD and MYOG were significantly reduced. This result illustrates that inhibitingHDAC2can improve the myoblast cell proliferation and reduces differentiation potential.The expression of HDAC2may play a role on inhibition of cell proliferationand promotecell differentiation during the development of myoblast cells.To sum up, four candidate genes are involved in the regulation of muscle growth.NCOA7, MYCN and HDAC2genes mainly involve in the functions of later embryonicdevelopment and growth before the28D.