Association Analysis Of Lactation Traits With The Bovine SCD1 Gene Promoter Activity

Milk fat is one of the main nutrients in milk,which plays a key role in affecting the quality and taste of milk.However,there is a phenomenon of low milk fat rate in China ’s dairy industry.How to improve milk fat rate has become an urgent problem to be solved.Studies have shown that SCD1 gene can directly affect the content of fatty acids in milk.In this study,the SNP loci in the promoter region of SCD1 gene were detected by direct Sanger sequencing,and the SNP loci significantly associated with lactation traits were identified.Then,the genotypes with positive correlation with milk fat rate were screened,and the effects of different genotypes on the promoter activity of SCD1 gene were further explored,so as to identify the genotypes that can promote the milk fat percentage.The results are as follows:1.Analysis of bovine SCD1 gene promoter activityThe transcription start site of bovine SCD1 gene was determined to be G(+1)by 5’-rapid-amplification of cDNA ends.The 1936 bp sequence of 5 ’-regulatory region of bovine SCD1 gene was obtained by PCR amplification.Then the fragment-deleted promoter sequences were obtained by subcloning and constructed into a dual luciferase reporter gene vector.After transfecting the successfully constructed recombinant plasmids into the bovine mammary epithelial cells,the double luciferase activity was detected.The results showed that the recombinant plasmid PGL3-570 /+111 had the highest enzyme activity,so the core promoter region of SCD1 gene was determined to be-570 to-1.2.Identification of SNP loci in the promoter region of bovine SCD1 gene and its association with lactation traitsThe promoter region of SCD1 gene in Holstein cows was detected by direct Sanger sequencing.The results showed that three SNP loci were present,which were g.21262844 G > A,g.21263552 A > G and g.21263651 G > A,respectively.Association analysis showed that only the SNP g.21263552 A > G in the core promoter region was significantly correlated with milk fat rate.The milk fat rate of AA genotype was significantly higher than that of AG genotype and GG genotype(P < 0.05).3.Effect of AA/GG genotype of SNP locus in core promoter region on promoter activityThe dual luciferase reporter gene vector of AA / GG genotype in the core promoter region was constructed,and then the dual luciferase activity was detected after transfection into the bovine mammary epithelial cells.The results showed that the promoter activity of GG genotype was significantly lower than that of AA genotype(P < 0.05).In conclusion,there was a significant correlation between the SNP locus g.21263552A>G in the core promoter region of the SCD1 gene identified in this study.The GG genotype significantly decreased the promoter activity of the SCD1 gene,which suggests that SNP locus g.21263552 A>G is significantly correlated with milk fat rate and could be utilized as a potential molecular marker for selecting cows with high milk fat rate.