ObjectiveMannose-binding lectin (MBL)-associated serine protease1 (MASP1) is the central enzyme in the innate immune system, which has biological functions of antibacterial and anti-inflammatory activities. Moreover, MASP1 represents a candidate gene reflecting the complement activity. This study is to investigate the entire exons of MASP1 in Chinese Holstein cattle with DNA sequencing to identify novel single nucleotide polymorphisms (SNPs). MethodsNovel SNPs were identified through gene sequencing and genotyped by the PCR Restriction Fragment Length Polymorphism (PCR-RFLP) and Created Restriction Site PCR (CRS-PCR). The relationship between the milk performance traits and complement activity in Chinese Holstein cattle was analyzed using the General Linear Model (GLM) procedure with the SAS software (version 8.0). ResultsTwo novel SNPs (i.e., g.5766A > G and g.51228A > C) were detected. The SNP g.5766A > G was located in the first intron and the SNP g.51228A > C was located in the 3′-untranslated regions of MASP1. The polymorphism at g.5766A > G was correlated with protein percentage (P < 0.05). Moreover, the polymorphism at g.51228A > C had only two genotypes, and this SNP had no significant correlation with CH50, ACH50, fat percentage, protein percentage, 305-day milk yields, or SCS scores. ConclusionMASP1, reflecting the complement activity, may not be significantly related to mastitis. However, MASP1 could be implemented in the breeding program to improve the production performance of Chinese Holstein cattle.