| Subgroup J Avian leukosis is composed of Subgroup J Avian leukosis virus(ALV-J)in clinic,is given priority to with hematopoietic cells malignant proliferation of a variety of tumor diseases.Since the 1990 s,it has seriously endangered China’s poultry industry and caused huge economic losses.Because of the international drug and commercialized vaccine for the prevention and treatment of J subgroup avian leucosis,it is only possible to achieve the control by eliminating positive chickens and strengthening biosafety measures.At present,the prevention of avian leucosis from host resistance is a hot research topic at home and abroad.ALV-J resistant chicken was found and screened in the early stage of the laboratory.On the basis of the experiment,the resistance of F1 generation chicken male female individual mating,the F2 chickling poison attack again,get the ALV-J resistant and susceptible chicken liver tissue samples Medip-Seq and RNA sequencing analysis.1.The results of Medip-Seq showed that,after comparison,there were 8304 differential methylation zones,among which the susceptible chicken(positive)was up to 4,709 compared with the resistant chicken(negative),and 3,595 were down-regulated.DMR enrichment analysis of different gene structures found that DMR had the largest concentration in the intergenic region,with 5,943,followed by introns.The promoter region had 233 different,accounting for 3% of the total.KEGG passway analysis of the difference gene showed that these differentially expressed genes were significantly enriched in the four signaling pathways of Steroid biosynthesis,Regulation of autophagy,beta-alanine metabolism,and ABC transporters.2.The RNA-Seq results showed that there were 515 different genes after screening,among which,there were 189 genes in susceptible chicken(positive)versus resistant chicken(negative),and 326 genes were down-regulated.KEGG passway analysis of the difference gene showed that these differentially expressed genes were significantly enriched in signaling pathways such as Glycosphingolipid biosynthesis,Protein processing in endoplasmic reticulum,Protein export,etc.3.According to the analysis results of Medip-Seq and RNA-Seq,the genes were randomly screened for BSP and qRT-PCR verification.It was showed a significant negative correlation between changes in the methylation level in promoter region and gene expression of TGFB2.The verification results proved the Medip-Seq and RNA-Seq data analysis result had higher accuracy and reliability.4.Medip-Seq data associated with RNA-Seq data analysis found that a total of 138 different gene methylation and expression,and had six gene who had a significant negative correlation between changes in the methylation level in promoter region and gene expression,including HSD17B7,SUPV3L1,TGFB2,TMEM104,FAM173 A,CNP1.So,we speculated that these six genes may be associated with ALV-J resistance.5.The expression of TGFB2 gene in DF-1 cells and HD11 cells showed an upward trend after infection with ALV-J.The proliferation of ALV-J in DF-1 cells was inhibited after si-RNA interference with the expression of TGFB2 gene.The expression of TGFB2 gene can affect the proliferation of ALV-J in DF-1 cells.It was speculated that TGFB2 gene promoter DNA methylation can affect the proliferation of ALV-J in resistant chicken by inhibiting the expression of TGFB2,and achieved the effect of disease resistance.So we guessed DNA methylation in the promoter region of TGFB2 gene could be used as a marker for screening of ALV-J resistant chicken. |
