| Laying traits are quantitative traits regulated by a complex genetic network.Chicken ovarian follicular development is regulated by a variety of genes.Both Wnt signaling pathways centered on CTNNB1 and miRNAs can participate in regulating ovarian follicular development.We found that the expression of CTNNB1 and mir-458b-5p in ovary of Jining Bairi chicken was different before and after the 140 day-laying,and the expression trend was opposite.It was further found that mir-458b-5p inhibited the proliferation of chicken grade granulosa cells in vitro by targeting CTNNB1.The purpose of this study was to study the association of chicken CTNNB1 and pri-miR-458 b polymorphic loci with egg laying traits.Using Jining Bairi chicken as the research material,the polymorphisms of CTNNB1 and pri-miR-458 b were found by mixed pool sequencing.The genetic loci are then typed by direct sequencing of the PCR products,and they are correlated with AFE and egg production,and it is expected that valuable molecular markers will be found.The main findings are as follows:(1)A total of 10 polymorphic loci were detected in the 5' regulatory region of CTNNB1.Four low polymorphic loci(-1295,-1249,-1154,-660,PIC < 0.25)were detected,and the remaining loci were all moderate polymorphism(0.25 < PIC <0.05).pri-miR-458 b detected 30 polymorphic sites and 6 moderate polymorphic sites(g.8954697,g.8954931,g.8954984,g.8955131,g.8955137,g.8955198),the other loci showed low polymorphism.(2)The 5' regulatory region of CTNNB1 at the locations-11060,-854,-817,and-782 are significantly related to egg laying traits.Compared with the CC type at the-1160 site,AFE was 3.33 days earlier,and the number of eggs produced by 18 to 23 weeks was about 3 eggs,and the total number of eggs was about 6 eggs.The combined genotype of the-1160 locus and the linkage locus significantly affected AFE and the number of eggs produced at 18 to 23 weeks.The-854 and-817 loci were linked,CA haplotype production started 3.89 days earlier,and the number of eggs laid earlier increased by 3.09 eggs.pri-miR-458 b has 10 loci that are related to egg production traits.Among them,g.8955225,g.8955339,g.8955341,g.8955348,g.8955384,and g.8955420 are linkage disequilibrium.The combined genotype significantly affected the number of eggs laid in the later period and the total number of eggs laid;the dominant haplotype AGCGCG increased the number of eggs laid by 3.29 and the total number of eggs laid increased by 8.08.(3)Using AliBaba2.1 online software to predict the transcription factor of the 5' regulatory region of CTNNB1,the transcription factor was predicted,and it was found that the transcription factors had changed before and after mutation at the four loci associated with egg production traits.Compared with the wild type,the types of mutant-bound transcription factors may increase or decrease.The-1160 site was able to bind to Sp1 and Oct-1 before mutation,and could only bind to Sp1 after mutation.The-854 mutation can not only bind to Sp1 and Egr-1,but also to Oct-1,NF-muE1,and NF-1.The-817 site can only bind to Sp1 before mutation,and can bind to Sp1 and Krox-20 after mutation.The-782 mutation can bind to EV-ErbA.It is speculated that these loci may affect egg laying traits by affecting the transcription of CTNNB1.In addition,methylation prediction revealed that a CpG island exists in the 5' regulatory region of CTNNB1,and three polymorphic sites(-854,-817,-782)that significantly affect egg production traits are located in the CpG island,indicating that these SNPs is possible to affect the egg production traits by changing the degree of methylation in this region.(4)Prediction of the secondary structure of different haplotypes of pri-miR-458 b,it was found that the free energy changes significantly,and the neck conformation of the stem-loop structure changes due to the base mismatch caused by the mutation.It is speculated that primiR-458 b mutations may affect the recognition and cleavage of endonucleases,thereby reducing the expression level of pre-miR-458 b and affecting the expression of mature miR-458b-5p.In summary,a total of 40 SNPs were found in the 5' regulatory region of CTNNB1 and pri-miR-458 b,and 14 polymorphic loci significantly affected the egg production traits of Jining Bairi chicken.The 5' regulatory region of CTNNB1 is linked at-1296,-1235,-11060 and-1154,and linked at-854 and-817.Their combined genotypes have a significant effect on AFE and the number of eggs laid in 18-23 weeks.pri-miR-458 b has 6 loci(g.8955225,g.8955339,g.8955341,g.8955348,g.8955384,g.8955420)in a linkage disequilibrium,and its combined genotype significantly affects late egg production and the total number of eggs laid. |
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