Screening The Genes Of Lipid Deposition In High-quality Meat-type Chickens And Their Expressions In The Process Of Preadipocytes During Differentiation

In order to study the the molecular mechanism of regulating fat deposition in the high quality chickens, we used AA broiler and high quality chicken HS1 line as experimental materials in this experiment to study the expression patterns of fat metabolism related genes at individual and cell level, expect to find out the candidate genes which regulate lipid metabolism, and provide scientific basis for further selection of high quality chicken.This experiment selected liver of AA and HS1 chickens at the embryonic period (21 d) for screening the differentially expressed genes between two chicken lines, especially fat metabolism related genes with gene expression profile analysis (DGE) method. We analyzed the differences genetic mechanism in fat metabolism between two lines from the level of transcriptome; Using Real-Time PCR quantitative analysis to determine the expression level of ACC, FAS, PGC-1α, PPARγ, SREBP-1c, and PLIN1 gene in AA and HS1 chickens at 28 d,49 d, and 70 d. We cultured the pre-adipocyte from abdominal fat of AA chickens at 12d, to analysis the expression regular of ACC, FAS, PGC-1α, PPARγ, SREBP-1c, and PLIN1 gene in the process of adipocyte differentiation. We proved the mechanism of candidate genes in the process of adipocyte differentiation on cellular level.Using AA and HS1 chickens liver to transcriptome sequencing by the DGE technology, we obtained 6.90-9.57 million original data from each library, and obtained 6.85-9.50 million clean data, identified 14977 genes. There are 1270 genes exist significant differences at P< 0.05 level and 493 genes exist differ significantly at P< 0.01 level. Through GO and Pathway analysis, these differences genes were enriched to biological processes such as cell apoptosis, cell cycle, gene enrichment sphingolipid metabolism, ether lipids metabolism, PPARs signaling pathways, fat cytokine signaling pathways, mTOR signaling pathway, and insulin signaling pathway. There are 34 lipid metabolism related genes exist significant difference between the AA and HS1 chickens, including PPARy and PLIN1. Fat metabolism related factor PPARy and PLIN1 expression in the liver of HS1 were respectively 3.79 and 5.42 times of AA chicken. In addition, SREBP-1c, FAS, ACC, and PGC-1a which closely related with PPARy also have differences between the two lines; the expression ratios between them were 0.29,0.76,0.17 and 0.61, respectively.Quantitative analysis candidate genes in liver, abdominal fat, and subcutaneous fat in AA and HS1 chickens, we observed varieties, age and organizational factors affected A CC, FAS, PGC-1α, PPARy, SREBP-1c, and PLIN1 gene expression, excepted for the fat synthesis related genes FAS. The other genes expression in the liver and abdominal fat of HS1 chickens were higher than AA broilers at 28 d,49 d, and 70 d; With the increase of age, the expression of ACC, FAS, PPARy, SREBP-1c, and PLIN1 in abdominal fat of HS1 chickens showed a trend of decreases after increase first. PGC-1α had an increasing trend of after decreases in liver, and showed a trend of rising in the abdominal fat of two chicken lines; PGC-1α in liver expressed higher than abdominal fat and subcutaneous fat. ACC, FAS, and SREBP-1c in abdominal fat expressed higher than liver and subcutaneous fat, PPARy and PLIN1 highly expressed in abdominal fat, and extremely low in liver and subcutaneous fat.In addition, this experiment had original generation of cultivation and differentiation the pre-adipocytes of chicken, CCK-8 method was used to detect the cell growth curve, found that the induction group and the control group pre-adipocytes begin to multiply in 36 h; the multiplication rate achieve maximum at 60 h; and cell proliferation rate decline at 72h; determination of fat content in cells by oil red O staining method, found that the induction group and the control group fat content in pre-adipocytes were lowest at 24 h, and began to increase after 24 h, reach the highest level at 60 h; significantly decreased at 72 h, and the fat content in induced group was obviously higher than that of control group after 24 h; quantitative analysis of ACC, FAS, PGC-1α, PPARγ, SREBP-1c, and PLIN1 in the process of adipocyte differentiation, found these genes highest expressed at 48 h, and expressed lower at 24 h and 70 h; expression in induced group were higher than the control group at 48 h and 70 h.Generally, in the present study, we screened differentially expressed genes in the liver of AA and HS1 chickens at embryonic period 21 d via gene expression profile analysis technology. The results indicated that 14977 genes expressed in both AA and HS1 chickens and 1270 of these genes expressed differently between AA and HS1. The factors of population, age, and tissue significantly affected the expression of ACC, FAS, PGC-1α, PPARy, SREBP-1c, and PLIN1 in the growth period of AA and HS1 chickens. In the process of adipocyte differentiation, the expression of ACC, SREBP-1c, and PLIN1 in inducing group were significantly higher than the control group at 48 h and 70 h, respectively, and the expression of FAS and PPARy in inducing group were significantly higher than the control group at 70 h. Thus we inferred that these genes can promote the process of adipocyte differentiation.