The Effects Of NCAPG On The Mitosis And Myogenic Differentiation Of Myoblasts Derived From Fetal Bovine Skeletal Muscle And Mechanism Study

Growth rate and carcass weight are the most important traits and influence the economic benefits in beef cattle industry.Therefore,it is of practical importance to cultivate breeds with rapid growth,high dressing percentage.Bovine skeletal muscle development initiates at the early pregnancy and mainly finishes in the late pregnancy.Under the influence of a variety of transcription factors,the development went through several stages including mesenchymal stem cell,myogenic progenitor,myoblast,primary fibers and secondary fibers.In order to explore the mechanism of growth rate candidate gene,NCAPG,in bovine development,we studied the function of NCAPG in skeletal muscle growth using in vitro model.The aim of this study is to investigate the function of NCAPG in the mitotic and differentiation of myoblast derived from the skeletal muscle of fetal bovine using RNA interference,and explore the functional mechanism.The main results were as follows:1 Myoblasts were isolated from skeletal muscle of three months bovine fetus using collagenase digestion.The myoblasts have strong differentiation ability.Myoblasts fused and formed amount of myotubes after inducing by horse serum culture medium.Myotubes were increased with the extension of time.The m RNA expression of MYH1,MYH2,MYH3 and MYH4 were upregulated significantly with the differentiation.While,the expression of MYOD and MYF5 were downregulated and the MYOG was steady expressed.2 Knocking down NCAPG with the siRNA in the stage of proliferation,the process of mitosis of myoblasts was prolonged about 25%.3 Knocking down NCAPG with the si RNA in the myoblast differentiation stage,myogenic differentiation related genes expression were significantly higher than negative control group at 1.5 day,while knockdown group were lower than negative control group at fourth day.4 Extracting the nucleus and cytoplasmic protein of the differentiation stage,we found that high amount of NCAPG existed in the nucleus,which implied that the NCAPG could contact chromatin and regulate the gene expression.5 Knocking down NCAPG with the si RNA,the K4K20me1 was decreased and the H4K16 ac was increased at the 1.5 day.This finding indicate that the chromatin become more unwound after the NCAPG knockdown.We speculate that there are some relationship between chromatin compaction and histone modification.6 Knocking down NCAPG with the siRNA,myoblasts of negative control group differentiated normally and form amount of myotubus at the fourth day,but myotubus of the knockdown group began to apoptosis at the fourth day and disappeared at the fifth day.