Study of Chicken Δ~6 Fatty Acid Desaturase Gene’s Promotor Region Polymorphism and Gene Expression
|School||Henan Agricultural University|
|Course||Animal Nutrition and Feed Science|
|Keywords||chicken FADS2 5’UTR SNP gene expression|
There is close relationship and interaction between nutrition and genetic factors. Nutrigenomics and nutrigenetics are very hot research. Fatty acid delta-6-desaturase is the rate-limiting enzyme in the synthesis of long-chain Polyunsaturated fatty acids（PUFAs）. PUFAs are very important for animals health, interrelated with fatty acid metabolism disease, thought as a important element which decided meat quality. But the gene structure is not fully understood, the gene function has not been a comprehensive study. In this research , firstly this gene’s 5’UTR has been cloned and precise transcription start site has been determined by using common PCR and 5’RACE. Secondly, using DNA pool sequencing , variable sites of chicken FADS2 gene’s promter region has been screened. And then SNPs of FADS2 gene’s promter region were genotyped in an F2 resource population of Gushi chickens crossing with Anka broilers using PCR-RFLP. The associations of SNPs polymorphisms with growth, carcass and meat quality traits, as well as sersum parameter in F2 resource were analyzed. Thirdly, Using real-time PCR techique, the mRNA of FADS2 gene in various tissues （heart, liver, crurreus） from male silky fowl （16 weeks old） were analyzed. The results as follow:1、The 5’RACE result showed that the cloned 196bp long 5’UTR region of chicken FADS2’gene, while it was found to extend 48 bp upstream to the known partial cDNA. The bioinformatics analysis revealed that a canonical TATA box and a GC box were present at the upstream of experimentally certified transcriptional start site （-28～-22 and -87～-79, respectively）. A typical CpG inland was found to stretch from the far upstream to the far downstream of transcriptional start site （-641～+200）. In addition, several copies of GATA-1/GATA-2, and SRY binding sites were predicted at the 5’flanking region.2、Nine SNPs were found in Chicken FADS2 gene’s promotor region. The two SNPs （C-958T,C-725T） in the F2 resource of the genetic polymorphisms were detected（AluⅠand HhaⅠ, respectively）.Population genetics analysised that: For C-958T locus, C、CC respectively as advantage alleles and advantage genotype. For F2 resource this locus was at Hardy Weinberg equilibrium. For C-725T locus, C、CC respectively as advantage alleles and advantage genotype. For F2 resource this locus wasn’t at Hardy Weinberg equilibrium. The C-958T locus was a moderate polymorphism, C-725T locus was a low polymorphism.The associations of C-958T、C-725T locuses polymorphisms with important traits in F2 resource were analyzed. The results showed as follow: C-958T locus was associated with body weight, Shank girth, Body slanting length, Chest width, Chest depth, Carcass weight, Head weight, Claw weight, Liver weight, Gizzard weight, Duodenum length, Aspartate Aminotransferase, Amylaceum and Glycerin Trilaurate.C-725T locus was associated with Heart weight rate, Gizzard weight rate, Leg muscle weight rate, Jejunum length, Ileum length, Alanine Transaminase, Total Cholesterol and C22:3、C22:4 in muscle.3、Analysis and comparison expression of silky fowl’s FADS2 in different tissue at different age（0 week-old,4week-old,8week-old,12week-old）. At 0 week-old, the expression of FADS2 in liver was significantly lower than the other time slice. There was was no significantly different of the expression of FADS2 in heart and crurreus. However, The expressions of FADS2 in liver was significantly higher than heart and crurreus in silky fowl （P < 0.05）,except at 0 week-old.From the above results, the conclusions can be as follow:1、Chicken FADS2 gene’s accurate transcription start site is at ATG upstream 196 bp. This gene’s promotor is a strong promotor.2、There are a lot of variation in chicken FADS2 gene promotor region. C-958T locus is significantly associated with many growth traits, carcass traits and sersum parameter. C-725T locus is significantly associated with a part of carcass traits , sersum parameter and PUFAs in muscle. It will be helpful for breeding as molecular markers.3、Silky fowl FADS2 gene expression showed temporal and spatial expression pattern. Liver is the major organs for metabolism of PUFAs. The silky fowl’s metabolism of PUFAs has been mature at 4 week-old.