Cloning, Expression Of FATP1/FATP4 Genes And Association Analysis Between Polymorphisms And Carcass Traits In Chicken

Long-chain fatty acids(LCFA) are both important metabolites and signaling molecules.At present,several candidate proteins have been proposed to be involved in the uptake process of LCFAs including fatty acid translocase(FAT/CD36),fatty acid binding protein(FABPs),long-chain fatty acyl-CoA synthetases(ACSL) and fatty acid transport proteins(FABPs).Although many of the above-mentioned proteins have important role in the proceeding of LCFAs uptake and metabolism,recent in vivo studies have particularly highlighted the contribution of FATPs to lipid metabolism and fatty acid-associated disorders.So,the studies about FATP are being focused in today because of FATP special character and biological function.In the studies,we adopted Sichuan Mountains Black-Bone chicken as materials. The code sequences of FATP4 protein were gained by bioinformatics analysis.The sequences were cloned with pMD18-T vector in E.coli DH5a and sequenced.Then, the basic physical parameters,glycosylation sites,phosphorylation sites, hydrophobicity and second structure were predicted by proteome analysis tools. Through analysis,we concluded that it was the protein of chicken FATP4.The present study also took Sichuan Mountainous Black-Bone chicken and S01 of Daheng chicken as materials.We used the real-time PCR(SYBR greenⅠ) method to test the expression quantity of FATP1 and FATP4 gene in different breed of above-mentioned at 10 week.And we tested the expression quantity of FATP1 and FATP4 gene in different tissues and the expression quantity at different growth points. The results showed that the expression quantity of FATP1 and FATP4 had no significant difference between the two breeds.Moreover,there was advantageous tissue for gene expression at each growth point and there was also advantageous growth point for gene expression in a certain tissue. The current study adopted eight different populations of Daheng high-quality chicken in Sichuan province as samples.Five SNPs in FATP1 gene were identified using PCR-SSCP method,including 49360bp(SNP1,G/A),48195 bp(SNP2,G/A), 46847bp(SNP3,A/G),46818bp(SNP4,A/G) and 46555bp(SNP5,A/G), respectively.Because the sample number of A46818G was few,we didn't analyze this mutation.The results of single locus analysis showed that at SNP1(G 49360A) locus, GG genotype had significantly higher(P＜0.05) breast muscle weight(BMW),leg muscle weight(LMW) and the percentage of breast muscle weight(BMWP) than AA and AG genotypes in 70d chicken populations;AG genotype had significantly higher live weight(LW),carcass weight(CW),semi-eviscerated weight(SEW), eviscerated weight(EW),LMW and the percentage of leg muscle weight(LMWP) than GG genotype in 91d chicken samples;AA genotype had significantly higher (P＜0.05) LW,CW,SEW and EW than AG and GG genotype,moreover,GG genotype had significantly higher BMWP than AA and AG(P＜0.05) in all examined samples.At SNP2(G48195A) locus,NN genotype had significantly higher(P＜0.05) BMW,LMW, BMWP and LMWP than MM and MN genotypes in 70d chicken populations;NN genotype had significantly higher LW,CW,SEW and EW than MM and MN genotype, while the NN genotype had significantly lower subcutaneous fat thickness(SFT) and the percentage of abdominal fat weight(AP) than that of MN genotype(P＜0.05) in 91d chicken samples;NN genotype had significantly higher(P＜0.05) LW,CW,SEW, EW,LMW,BMWP and LMWP than MM and MN genotype in all samples.At SNP 3 (A46847G) locus,in 70d and 91d chicken populations,there was no significantly association between genotype and carcass traits(P＞0.05);while AA genotype had significantly higher LW and CW than AG genotypes(P＜0.05) in all samples.At SNP5 (A46555G) locus,in 70d chicken populations,MN genotype had significantly higher (P＜0.05) LW,CW and EW than MM genotypes,but had lower LMWP when compared with NN genotype;in 91d chicken populations,NN genotype had significantly higher LW,CW,SEW,EW and LMWP than MM and MN genotypes, while the NN genotype had significantly lower BMWP than that of MN genotype (P＜0.05);in all chicken populations,NN genotype had significantly higher(P＜0.05) LMWP than MM and MN genotypes.The haplotype analysis indicated that in 70d chicken populations,the haplotype combinations had significant genetic effects on LW, CW,EW,BMW,LMW,SFT,BMWP and LMWP(P＜0.05);in 91d chicken populations,the haplotype had significant genetic effects on LW,CW,SEW,EW,SFT, LMWP and AP(P＜0.05);in all chicken populations,the haplotype had significant genetic effects on LW,CW,SEW,EW,SFT,BMWP and LMWP.Eight SNPs were detected at 5108778bp(G/A),5108695bp(G/A),5108287bp (C/-),5107545bp(T/C),5106740bp(C/T),5106005bp(A/G),5105481bp(G/A) and 5105454bp(C/T) in FATP4 gene.The allele A,C,C and A were the predominate allele in SNP2,SNP3,SNP5 and SNP6,respectively.The single locus analysis indicated that single nucleotide polymorphisms at SNP1(G5108778A) locus were significantly associated with BMWP and LMWP(P＜0.05) in 70d chicken populations;SNP1 was associated significantly with BMWP and AP(P＜0.05) in 91d chicken populations; SNP 1 was associated significantly with LW,CW,SEW,EW,AW,BMWP,LMWP and SFT(P＜0.05) in all samples.At SNP2(G5108695A) locus,there was no significantly association between genotypes and carcass traits(P＞0.05) in 70d and 91d chicken populations;while AW and SFT of the GG genotype were significantly higher than those with the AG genotype(P＜0.05) in all samples.At SNP3(C5108287-) locus, genotypes at this locus were significantly associated with AW,LMW-P and AP(P＜0.05) in 70d chicken populations;while,the TT genotype had higher BMWP than that of CT (P＜0.05) in 91d chicken populations;TT genotype had significantly higher AW, BMWP,LMWP and AP than CT and CC genotype(P＜0.05) in all chicken populations. At SNP4(T5107545C) locus,TT genotype had significantly higher(P＜0.05) BMW, LMW and LMWP than CC genotypes in 70d chicken populations,moreover,the SFT of chicken with the TC genotype was significantly higher than that of CC genotype; TT genotype had significantly higher LW,CW,SEW and EW than CC genotype in 91d chicken populations;TC genotype had significantly higher(P＜0.05) LW,CW and EW than TT genotype,moreover,TT genotype had significantly higher LMWP than CC(P＜0.05) in all chicken populations.At SNP5(C5106740T) locus,SNP at this locus was only significantly associated with BMWP(p＜0.05) in 70d chicken populations;in all chicken populations,single nucleotide polymorphisms were significantly associated with SFT(p＜0.05).At SNP 6(A5106005G) locus,AG genotype had significantly higher BMW and BMWP than AA and AG genotype (P＜0.05) in 70d chicken populations;GG genotype had significantly higher LW,CW, SEW,EW and LMW than AA genotypes(P＜0.05) in 91d chicken populations.At SNP7(G5105481A) locus,GG genotype had significantly higher(P＜0.05) LW and EW than AG genotypes in both 70d and 91d chicken populations;in all chicken populations,AG genotype had significantly higher(P＜0.05) BMWP than AA genotype. At SNP8(C5105454T) locus,there were no significant difference among all samples (P＞0.05).The haplotype analysis indicated that in 70d chicken populations,the haplotype combinations had significant genetic effects on LW,CW,EW,BMW,LMW, AP,BMWP and LMWP(P＜0.05);in 91d chicken populations,the haplotype had significant genetic effects on LW,CW,SEW,EW,AW,SFT,BMWP,LMWP and AP (P＜0.05);in all chicken populations,the haplotype had significant genetic effects on LW,CW,SEW,EW,LMW,AW,SFT,BMWP,LMWP and AP(P＜0.05).