Identification of candidate genes responsible for innate fear behavior in the chicken

Identifying the genes responsible for quantitative traits remains a major challenge. We previously found a major QTL on chromosome 4 affecting several innate fear behavioral traits obtained by an open-field test in an F2 population between White Leghorn and Nagoya breeds of chickens (Gallus gallus). Here, an integrated approach of transcriptome, haplotype frequency, and association analyses was used to identify candidate genes for the QTL in phenotypically extreme individuals selected from the same segregating F2 population as that used in the initial QTL analysis. QTL mapping for the first principal component, which summarizes the variances of all affected behavioral traits in the F2 population, revealed the behavioral QTL located at 14-35 Mb on chromosome 4 with 333 genes. After RNA-seq analysis using two pooled RNAs from extreme F2 individuals, real-time qPCR analysis in the two parental breeds and their F1 individuals greatly reduced the number of candidate genes in the QTL interval from 333 to 16 genes. Haplotype frequency analysis in the two extreme F2 groups further reduced the number of candidate genes from 16 to 11. After comparing gene expression in the two extreme groups, a conditional correlation analysis of diplotypes between gene expression and phenotype of extreme individuals revealed that NPY5R and LOC101749214 genes were strong candidate genes for innate fear behavior. This study illustrates how the integrated approach can identify candidate genes more rapidly than fine mapping of the initial QTL interval and provides new information for studying the genetic basis of innate fear behavior in chickens.