Improving the production traits and resistance against mastitis in dairy cattle is a challenge for animal scientists across the globe. The present study was designed to investigate the genetic effects of single nucleotide polymorphisms (SNPs) in Janus kinase 2 (JAK2) and diacylglycerol acyltransferase (DGAT1) genes with production and mastitis-related traits. Four SNPs in JAK2 and one in DGAT1 were analyzed through Chinese Cow's SNPs Chip-I (CCSC-I) and genotyped in a population of 312 Chinese Holsteins. Our findings demonstrated that milk fat percentage, somatic cell count (SCC), somatic cell score (SCS), serum cytokines interleukin 6 (IL-6) and interferon gamma (IFN-γ) showed significant associations (P < 0.05) with at least one or more identified SNPs. Consequently, the analysis based on haplotypes amongst the SNPs in JAK2 revealed noteworthy (P < 0.05) association with SCC and IL-6. Collectively, our results verified the pleiotropic ability of detected SNPs in bovine JAK2 and DGAT1 for milk fat percentage as well as mastitis-related traits. The significant SNPs in both the genes could serve as powerful genetic markers to minimize mastitis risk. In addition, besides SCC and SCS, the IFN-γ and IL-6 could also be used as indicators of improved genetic resistance against mastitis.