Polymorphisns In The5’ Region Of Chicken FADS2Gene And The Promoter Function Analysis

Delta-6fatty acid desaturases, which encoded by the FADS2gene, is arate-limiting desaturase in the biological metabolic process of fatty acid, is a member ofthe fatty acid desaturase (FADS) gene family. Polyunsaturated fatty acids is the importantcomposition of the cell membrane fluidity, and regulate lipid metabolism, promote animalgrowth, improve animal reproductive performance, improve the flavor of the meat. Forbirds, the content of polyunsaturated fatty acids directly affects the flavor.We used DNA pool sequencing method to scan the SNP sites of the FADS2gene andPCR-RFLP and CRS-PCR-RFLP technic to analyse the SNP distribution and populationgenetic characters in Gushi Anka F2resource population.9SNPs were found in the5’region of the FADS2gene. Linkage disequilibrium was analyzed between the SNP sites.Haplotypes were constructed based on haplotype structure of the SNPs in all experimentalanimals. Associations between SNPs and broiler growth traits, cacarss traits, meat quality,serum biochemical parameters and fatty acid contents in the muscle were determined aswell as the additive and dominace effect of the two SNP sites on the related traits. Weconstruct a series of recombinant plasmids with different length segments of the FADS2gene promoter regions, in order to study the gene transcription regulation. Transfectioncells, dual luciferase activity analysis were used to study the expression of the gene activity,and to determine the core promoter sequenceof the FADS2gene. The main results are asfollows:(1) The SNPs of C-1757T and C-1402T were in low polymorphism, with only2genotypes. The C-1757site had significant associations with the body weight of4,8,10,12weeks, Shank length of0,8,12weeks，body length and breastbone length of12weeks, Carcass weight, breast muscle weight and Leg muscle weight (P <0.05). At thesame time, C-1757site had significant associations with High Density Lipoprotein and LowDensity Lipoprotein (P <0.05). C-1402T polymorphism had significant associations withthe Birth weight and body weight of2,4weeks (P <0.05).(2) The G-997C polymorphism showed significant associations with BW2, BW4,BW6, SL4, SG4, BBL4, α-linolenic acid (C18:3) and arachidonic acid (C20:4)(P <0.05).The mean values of these traits were significantly higher in genotype CC than genotypesCA and AA. For the traits of BW4, BW6, BBL4and linoleic acid (C18:2), the additiveeffects were significant (P <0.05), and the dominance effect was significant (P <0.05) for arachidonic acid (C20:4). Significant associations for the A-996C polymorphism werefound for Birth body weight (BBW), BW2, BW4, BW6, SG4, BBL4, linoleic acid (C18:2)and arachidonic acid (C20:4) in this study (P <0.05). Besides the chickens with GCgenotype had the higher SL4, for other traits, GG genotype was significantly higher thanGC and CC genotypes. In testing for additive and dominant effect, the additive effects weresignificant (P <0.05) for BBL4and C18:3, and the dominance effect was significant (P<0.05) for arachidonic acid (C20:4). Besides, two SNPs had significant associations withγ-Glutamyl Transpeptidans (P <0.05).(3) The two SNPs were in strong LD, and it was estimated as D’=0.975, r2=0.459.Through estimating SNP haplotype frequencies, three haplotypes were observed in theindividuals of the F2resource population, including H1(C-A), H2(C-C)and H4(G-C).There existed six haplotype combinations in the F2resource population. Significantassociations were found between the haplotype combinations and linoleic acid (C18:2),α-linolenic acid (C18:3), arachidonic acid (C20:4), BW2, BW4, BW6, SG4and BBL4(P<0.05).(4) A series of different length fragments of chicken FADS2gene luciferase reporterplasmids with the same3’ terminal was successfully constructed, the plasmids ofPGL4.10-100and PGL4.10-200had a lower transcriptional activity. The activity ofPGL4.10-300was higher than PGL4.10-100and PGL4.10-200. The activity ofPGL4.10-500, PGL4.10-800and PGL4.10-1100were significantly higher thanaforementioned three plasmids (P <0.05).Conclusions:(1) The results suggested that the SNPs ofG-997C和C-1402T in the5’region ofFADS2gene affected the content of essential fatty acid in muscle, and played a role in theearly-stage growth rate of chickens.(2) The core promoter of chicken FADS2gene located at-319bp~-28bp, and anegative transcription factors may exist in the region of-1269~-902bp.