Fine Mapping The QTL For Growth Traits In Chicken By Improved Genotyping By Sequencing

Growth traits is one of the most important ecnomic trait in chicken.For decades,genetic improvement based on conventional breeding approach has been successfully applied in broilars.However,people are still lack of the knowledge on the genetic basis of the traits.Economically important traits in domestic animals are generally complex.Most of the complex traits are determined by both genetic and environmental effectors.Great efforts have been made to deciphering the genetic architecture of complex traits in past dadecades,but the task still be extremely challenging.Quantitative trait locus(QTL)mapping in animal populations has been a successful strategy for identifying genomic regions that play a role in complex diseases and traits.But due to the reasons of extensive linkage disequilibrium,limited genetic markers and interfering by multigene and small effects,the QTL interval is frequently large,often encompassing hundreds of genes.To narrow the locus and identify candidate genes,additional strategies are needed.Many studies have confirmed that there was a major effect QTL affects body weight at the distal end of chicken chromosome 1(165-175 Mb).In our previous research,QTL analysis in a reciprocal F2 cross between the Huiyang Beard chicken(HX)and the fast growing commercial broiler breed(A03)also revealed a growth QTL in the same region on chromosome 1.To physically reduce the chromosomal interval and the number of potential candidate genes,we performed fine mapping using individuals of generations F9 and F12 in an advanced intercross line(AIL)that established from the initial F2 mapping population.To achieving an optimal balance between sequencing resolution and budgets,especially in large-scale population genetics research,we established an improved double-enzyme digestion genotyping by sequencing(ddGBS)method on chicken.We evaluated eight double-enzyme digestion combinations ultizing five paramaters,and EcoRⅠ-MseⅠ was confirmed to be the optimal combination for the chicken genome.A total of 291,772 high-density SNPs from F9 generation individual animals were identified.We found that the consistency between ddGBS data and the Chicken 60K SNP Array is over 99%.The approach can greatly benefit the fine-mapping of the QTL and identification of causative genes.GWAS analysis for F9 generation animals reconfirmed the previously mapped growth QTL on chromosome 1(Chr1:168.2-171.3 Mb).This QTL affects 21 growth traits such as body weight at 2-14 weeks of age and length of intestine.The peak obtained with the AIL was narrower than with the original F2 population.Meanwhile,we identified a narrow region(Chr27:3.60-3.75 Mb)to be strongly associated with shank length at 6-12 weeks of age.Haplotype-sharing approach was used to refine the map position of the QTL.We genotyped additional SNP markers in this region that were nearly fixed in different directions in the two F0 lines to assign their parental origin in F9 individuals.IBD(Identical-by-descent)analysis identified four recombination breakpoints that divided the QTL interval into five haplotype blocks and we further narrowed the major QTL down to the Chrl:168.6-169.8 Mb interval.Unfortunately,this region shows a mosaic pattern of association.We found two shared linkage regions among three broiler breeds based on the haplotype analysis of nine outgroup populations,and confirmed the significant correlation with body weight in F9 generation.In addition,differentially expressed analysis in this QTL region was performed between high and low body weight individuals in F12 generation.The expression of MLNR has significant difference in duodenum and proventriculus.We also found 365 genes which were located in whole genome potential QTL regions were significantly enriched in the neuroactive ligand-receptor interaction pathway(including MLNR and various kinds of growth hormone).For motilin is a key regulatory hormone for appetite,we consider MLNR as a functional candidate gene for body weight.In summary,based on the improved ddGBS method,serveal genetics approches were employed to fine map the QTL associated with growth traits.We expound the associated model of complex quantitative traits and provide new evidence for the genetic mechanism for further studies on growth related traits.