Regulation Of MYL4 Gene On Pig Muscle Growth And Development

The animal body is mainly composed of muscle tissue,and the growth rate and quality of muscle are very important economic characters in animal production.Muscle growth and development is an extremely complex process that can be affected by many aspects.Therefore,it is of great theoretical and practical significance to find the key factors regulating the growth and development of pig muscle.Early studies on myosin light chain 4 gene(MYL4)mostly focused on cardiomyopathy,but with the continuous in-depth understanding of MYL4 gene,it was found to be closely related to the growth and development of skeletal muscle.At the same time,based on the combined analysis of proteomics and transcriptomic,MYL4 gene was identified as a key candidate gene for pig growth and development.However,it is not clear how MYL4 gene is involved in the regulation of pig growth and development.In order to explore how MYL4 gene is involved in the regulation mechanism of growth and development of pigs,this study selected the small pig breeds with local characteristics--Tibetan pig and the fast-growing pig breed introduced from abroad--Yuoku pig as the experimental research objects,and cloned pig MYL4 gene by homologous recombination technology and carried out related bioinformatics analysis.With functional verification of porcine muscle growing-related genes as the main direction,the expression difference and gene function of MYL4 gene were explored at m RNA level and cellular level by overexpression and interference techniques,and the regulatory mechanism of MYL4 gene was clarified.The results are as follows:1.Semi-quantitative PCR was used to detect the expression levels of MYL4 gene in the heart,liver,spleen,lung,lung,kidney,dorsal fat,abdominal fat,longissimus dorsi,leg muscle and loin tissues of Tibetan pigs and about gram pigs,and to construct the expression profile of porcine MYL4 gene tissue.Real-time quantitative PCR was used to detect the relative expression of MYL4 gene in Tibetan pigs and gram pigs.The results showed that the expression level of MYL4 gene was the highest in liver and lung tissues of approximately gram pigs,followed by that in muscle tissues of large York pigs was higher than that of Tibetan pigs.In longissimus dorsi muscle tissue,the m RNA expression of MYL4 gene of approximately gram pigs was significantly higher than that of Tibetan pigs(P < 0.05).2.The vector of MYL4 gene was constructed by homologous recombination technique,and the full-length CDS region of porcine MYL4 gene was cloned successfully.Bioinformatic analysis of the sequence was conducted on the online website,and biological information such as secondary structure,tertiary structure,phosphorylation site and hydrophobicity of 197 amino acids encoded by the gene were analyzed.Results The molecular weight of MYL4 protein was 21 606.64 u.The molecular formula is C961H1514N252O295S9;Fat coefficient was 67.97;Is the hydrophilic protein,the theoretical isoelectric point p I is 4.69;The instability coefficient was 58.38,indicating unstable physical and chemical properties.The nucleotide sequences of CDS region of Tibetan pig MYL4 gene were 100 %,93.3 %,93.2 %,93.0 %,91.9 %,91.2 %,87.3%,81.5 % and 72.7 %,respectively,between wild boar,cow,dog,sheep,Homo sapiens,horse,mouse,chicken and zebrafish.3.In this study,MYL4 gene overexpression vector was successfully constructed,and si RNA with optimal interference efficiency was screened and transfected into Porky skeletal muscle satellite cells(PSMSCS),and the regulation of MYL4 gene on Porky skeletal muscle satellite cells was verified by EDU staining proliferation experiment.The influence of overexpression and interference of MYL4 gene on muscle hypertrophy and degradation marker genes in PSMSCS was also detected.The results showed that 48 h after transfection,overexpression of MYL4 gene could promote the proliferation of muscle cells,and interference with MYL4 gene could inhibit the proliferation of muscle cells.Moreover,MYL4 gene can promote muscle hypertrophy and inhibit muscle degeneration.In summary,the experimental results show that MYL4 gene is involved in regulating the regulatory base mechanism of pig muscle growth and development,and MYL4 gene is a key regulatory factor regulating pig muscle growth and development.The results of this study provide a certain theoretical basis for exploring pig muscle growth and development.