Mapping Inhibitor Of Dermal Melanin And Identification Of Markers Assisted Of Related Qualitative Traits In Chicken

To satisfy the increasing demands for livestock and poultry meat quality, improvement of meat quality has became a subject of increasing research. In addition to the requirements of quality of broiler meat quality itself nutrient content, flavor and taste, and so on, high quality chicken also need to have specific properties of packing. On the one hand, packaging characters can play the role of anti fake mark, on the other hand, it can also meet the special preferences of different areas of consumer. For poultry, quality traits associated with pigment had shank color and skin color. In order to establish molecular marker assisted selection(MSA) technology related melanin packaging traits in quality broiler breeding, this paper aim at mapping inhibitor of dermal melanin(Id) gene by targeted next-generation sequencing based on bulked segregant analysis(BSA),developing the molecular marker of willow shank, black skin and white feather, analyzing the polymorphic of the three loci, to set up molecular marker assisted selection system.This study obtained the following main results:Experiment 1 Mapping inhibitor of dermal melanin by targeted next-generation sequencing in chickenUsing of F2 resource population of Gushi chicken and Anka chicken, we establish the shank color extreme phenotype mixing pool-- yellow shank DNA pooling and willow shank DNA pooling, and conducted 200× deep sequencing at the 10 Mb interval with Chr Z 67.1-72.3 Mb as the core region, on the two pools by targeted next-generation sequencing at target region. By SHOREmap and differences mutation sites analysis of the data of yellow shank DNA pooling and willow shank DNA pooling, we mapped the inhibitor of dermal melanin(Id) gene at the interval for 71.58-72.18 Mb of chromosome Z in chicken, which reducing the interval of inhibitor of dermal melanin gene, and laying the foundation for the cause mutation of willow shank in chicken.Experiment 2 Identified candidate genes of inhibitor of dermal melanin in chickenThe differences analysis of 31 homozygous mutations obtained, we choose 6 mutations(according by the location of mutations, for example located in exon, intron, gene regulatory region and its neighborhood), and named the site I – VI, respectively. The correlation analysis of genotype and shank color in the chicken, which including for F2 resource group and chicken with different shank color(Gushi chickens, Silkies, the sea Department) and F1 generation of the pros and cons of Silky chicken and Hisex A line.The results found that the genotype of loci III-V was significantly associated with shankcolor.The position of the three loci on chromosomes was located in the intron of CDKN2 A gene,upstream of CDKN2 B gene, and the intergenic between CDKN2 B gene and another gene,respectively. According to the characteristics of genotype method, developing site IV as molecules for willow shank locus genotype. We analysis of candidate gene(MTAP,CDKN2 A and CDKN2B) expression in chicken from different shank color, and the mutation analysis in bioinformatics. According to the results of linkage analysis,expression of tissue analysis and biological information analysis, we think that the CDKN2A/B gene is the candidate gene of inhibitor of dermal melanin gene in chicken.Experiment 3 Verification causative mutation and development of the molecular markers of black skin in chickenThe results of copy number variation of Chinese of local chicken by our group found that the Silkies chicken and Xichuan chicken both has a copy number variation of CNVR(Chr 20 10717668 – 10845246 bp and 11265289- 11435353 bp). Using QPCR technology, we validated the results of copy number variation by a CGH. Meanwhile, the result of QPCR found the copy number variation in Fm loci of Silky chicken was about 2,and the result of QPCR found the copy number variation in Fm loci of Xichuan chicken was about 1.5-2. In order to explore the differences of the copy number variation in Fm loci between Silky chicken and Xichuan chicken, we obtained the same result after expanding the validation group. In order to establish the method of determining the Fm genotype, we determined the copy number variation of Fm loci of the individual coming from F1 generation of the pros and cons of Silky chicken and Hisex A line. The results shows that if the copy number at Fm loci of fmfm genotype was 1, Fmfm genotype was approximately 1.5, Fm Fm genotype was approximately 2. The molecular markers and genotyping methods can be used for identification homozygous in black skin site in chicken, and can provides a theoretical basis for marker assisted selection in black skin chicken breeding.In summary, this study mapped the inhibitor dermal melanin gene between71.58-72.18 Mb in chromosome Z of chicken, and points out the CDKN2A/B gene might be the candidate gene of inhibitor of dermal melanin(Id) gene in chicken, which reduce the positioning interval of Id gene in chicken, and lays a theoretical foundation for revealing the forming mechanism of shank color. This study also developed molecular markers linked willow shank and black skin traits, and developed molecular marker of Id and Fm locus, providing the theoretical basis and thinking for marker assisted selection breeding in quality traits associated with melanin – willow shank and black skin.