Cloning And Expression Character Analysis Of Akirin2 And MiR-155 From Different Strains Of Chicken

Screening and identification of genes related to muscle development and disease resistance have important theory references and utility values for chicken breed improvement and breeding. NF-κB is an important transcription factor for the regulation of immune response and the differentiation of muscle cells. AKIRIN2 positively regulates transcription of a variety of genes including IL-6 through indirect interacting with NF-κB. Moreover, akirin2 is related to intramuscular fat content of cattle and development of avian muscle fiber. miR-155 is a non-coding small RNA which is regulated by NF-κB. miR-155 is not only involved in the regulation of myeloid cell proliferation, activation and T, B cell differentiation, but also can regulate muscle cell differentiation through targeting the genes related to muscle development. This study selected muscle development and immune related target genes akirin2 and miR-155 and analyzed its sequence information and expression distribution characteristics in different strains of chicken. We hope it can provide theoretical references for screening candidate genes related to chicken strain improvement and molecular breeding. The main research methods and results are as follows:The experimental results of immunization with Newcastle disease vaccine in chicken showed that akirin2 had high expression level in the liver, but no obvious changes in other tissues on seventh day after immunization. However, akirin2 had high expression level in the bursa and low expression level in the liver on fourteenth day after immunization, respectively. Research results implied that akirin2 was not only involved in the innate immune response, but also played important roles in adaptive immunity.Sequence comparison and expression analysis of akirin2 between different chicken strains showed that 4 SNPs existed in 5’ terminals of akirin2 coding regions from the four strains of chicken, and which resulted in three amino acid changes of AKIRIN2 protein. The prediction results of protein secondary structure showed that there were no differences among the three strains of chicken. Their alpha helixes and random coils were formed in the same positions, but several protein binding sites existed subtle changes in quantities and location, speculating that these changes might produce some affections on the expression and activity of AKIRIN2 protein. The expression analysis of akirin2 gene in the different days old chicken of the same strain(Sanhuang broiler) showed that akirin2 were widely expressed in different tissues. The high expression existed in immune related tissues, such as bursa, spleen and blood. With the increasing of age, the expression of akirin2 in muscle showed a trend of decreasing. The expression analysis of akirin2 gene in different strain chicken of the same days old(Sanhuang and AA+ broiler) showed that the expression levels of akirin2 were lower in the heart, muscle, muscle stomach and thymus, and the higher levels of expression were in the blood, liver, large intestine and glandular stomach. However, the expression levels of the fat and spleen in the two chicken strains were different.The results that sequence analysis and tissue expression profiles showed that pri-mi R-155 from three strains of chicken had the same length without base mutations, which implied that processing and modification mechanisms of miR-155 were consistent in the three chicken strains. Phylogenetic analysis showed that mature miR-155 was highly conserved in each class vertebrates and homology was up to 100%. The homology of pre-mi R-155 is lower than that of the mature miR-155. Tissue expression profile analysis showed that mi R-155 of the three chicken strains were widely expressed in different tissues and no tissue specificity, but the high expression existed in immune related tissues and the low expression existed in muscle related tissues, respectively.Through cloning, expression character, and functional analysis of akirin2 gene and miR-155 from different chicken strains, the research results not only lay the foundations for further studying the functions of these genes, also provide references for the in-depth study of poultry breeding, immune and muscle development.