Studies On Genetic Effects Of DDIT3,RPL23A,SESN2 And NR4A1 Genes On Milk Production Traits In Chinese Holstein

Our previous RNA-seq study with the mammary glands of four lactating Holstein cows with extremely high and low phenotypic values of milk protein and fat percentages identified 31 candidate genes,Out of them,in this study,we selected four candidate genes,including DDIT3?RPL23A?SESN2 and NR4A1,to investigate whether they have significant genetic effects on on milk production traits in Chinese Holstein.Through re-sequencing the entire coding regions and 3000 bp of both upstream and downstream regulatory regions of the four genes by using the pooled genomic DNA of 40 Holstein bulls,a total of 35 SNPs and three indels were identified,including three in DDIT3,12 in RPL23A,16 in SESN2,and seven in NR4A1;moreover,two indels,g.125714860 AGCGGGGTGGGGG>-and g.125714806 CCCC>A in SESN2 were novel and have not been reported previously.A total of 1093 Chinese Holstein cows were genotyped for these identified SNPs using MALDI-TOF-MS.Subsequent single SNP analyses revealed multiple significant association to all 35 SNPs and three indels regressed against the dairy production traits(P<0.0001-0.0493).In addition,with linkage disequilibrium(LD)analysis,we found one,one,three,and one haplotype blocks in DDIT3,RPL23A,SESN2,and NR4A1 genes,respectively.Haplotype-based association analyses revealed some haplotypes were also significantly associated with milk production traits(P ? 0.0001?0.0461).We predicted the impacts of the alleles on each SNP in the 5'promoter region of DDIT3,RPL23A,SESN2,and NR4A1 genes on potential binding sites using JASPAR software,and found that two,two,nine and one SNPs changed the transcription factor binding sites on the promoter region of the above four genes,respectively.We also found that 11 SNPs and three indels(two in DDIT3,two in RPL23A,nine in SESN2,and one in NR4A1)altered the specific transcription factor binding sites(TFBS)in the promoter thereby regulating the promoter activity,suggesting that they might be the promising potential functional mutations for the milk traits.Together,our findings first indicated the DDIT3,RPL23A,SESN2,and NR4A1 genes had significant genetic effects on milk production traits in Chinese Holstein and verified the results of our previous RNA-seq study.Several SNPs,including g.56283814 C>T and g.56284880 C>T in DDIT3 gene,g.20702122 C>G in RPL23A gene,g.125714723 A>G,g.125714806 CCCC>A,g.125716120 C>T,and g.125716884 A>G in SESN2 gene,and g.27992897 C>T in NR4A1 gene,are potential regulatory mutations for the exprsssion of the four genes and are needed to perform in-depth validation investigation.