MBL1 Gene Polymorphisms And Its Association With Milk Production Traits And Mastitis In Bovine

This study analysis single nucleotide polymorphisms of mannose-combining lectin gene in bovine using molecular biology, and associations with milk performance traits, MBL-A level of serum and complement activity (CH50 and ACH50), inspection MBL-A antibacterial activity and MBL1 gene expression in the organization. For Chinese holstein cow with high-yielding breeding and resistance to provide certain theoretical basis and material. The results were as followed:1. In this study, three single nucleotide polymorphisms (SNPs) in promoter and two SNPs in exonl of MBL1 gene were detected by PCR-SSCP, PCR-RFLP, CRS-PCR, Nested-PCR and DNA sequencing methods in 537 individuals from three Chinese cattle breeds. The dominant alleles in 3 three breeds were A,C,G and G, and the allelic frequencies are A (CH0.9909 /LYC0.7 273/BBC0.9375),C (CH0.6 098/LYC0.7 841/BBC0.6250),G (CH0.7111 /LYC0.8523/BBC0.8958)和G (CH0.6130/LYC0.7614/0.7500). Theχ2 test for the Bohai Black Cattle of all sites achieved balance about the Hardy-Weinberg equilibrium (P> 0.05); Holstein cow in the mutation g.-2194 G>A has achieved Hardy-Weinberg equilibrium (P> 0.05); Luxi cattle only in the g.105 T>C sites has not achieved Hardy-Weinberg equilibrium (P＜0.05)2. Furthermore, analyzing the presence of the genotypes and haplotypes, we show the polymorphisms and their possible implications, while attempting to define the association of discovered polymorphisms with the serum MBL-A levels, complement activity (CH50 and ACH50) and the milk production traits. Homozygous genotype AA had significant lower than those with genotypes GA and AA(P<0.05) for CH50 in g.-1330 G>A. At g.70 G>A, the homozygous genotype GG had significant lower than those with genotypes GA and AA(P<0.05) for concentration of serum; genotype AA hah significant higher CH50 than genotype GG(P<0.05). However, no other significant associations were observed g.-2194 A>C, g.-1 446 and g.105 T>C polymorphisms and milk production traits and concentration of serum,CH50 and ACH50 in the 469 Chinese Holstein population (P>0.05).Statistical analyses revealed that cows with the combined genotypes of H16H17 had significantly lower SCS than combined genotypes H8H21The individuals with haplotype combination H14H1(P<0.05) presented a higher 305d milk yield than the ones with haplotype combination H16H1, H16H16, H16H17, H16H21, H16H3, H16H5, H16H6, H16H8, H8H1, H8H22 and H8H5, respectively;For fat percentage, the individuals with haplotype combination H16H7 (P＜0.05) showed significantly lower contrast to the haplotype combination H14H1,H16H16, H6H1 and H8H1; In protein percentage, the individuals with haplotype combination H16H21 (P＜0.05) and H5H1 (P＜0.05) presented higher than the ones with haplotype combination H14H1, H14H5,H15H1, H16H17, H16H22 and H8H17; For concentration of serum, the individuals with haplotype combination H7H3(P＜0.05) showed significantly lower contrast to the haplotype combination H14H5, H15H1, H16H1, H16H13, H16H15, H16H16, H16H17, H16H21, H16H3, H16H5, H16H6, H6H1, H8H1, H8H17 and H8H5; In the CH50, the individuals with haplotype combination H7H3 (P＜0.05) showed significantly lower than the haplotype combination H14H5, H15H1, H16H1, H16H21, H16H22, H16H3, H16H5, H16H7, H8H1, H8H21, H8H22 and H8H5; The individuals with haplotype combination H16H16 (P＜0.05) showed significantly higher ACH50 in contrast to the haplotype combination H16H21, H16H3, H16H5, H5H1 and H8H1.3. To probe the biological activity of MBL-A influence Staphylococcus Aureus and Escherichia coli, we counted the CFU. The results shown that serum from H16H17(resistant) animals displayed a significantly(P<0.05) higher antibacterial activity(Staphylococcus Aureus and E.coli infection) compared with serum from H8H21 (susceptible) animals, serum samples pre-incubated with mannose and serum samples pre-incubated with N-acetyl-D-glucosamine. The evidence that genetic analysis and functional (serum antibacterial activity) analysis yield fully concordant results lends biological significance to the protective role of the H16H17 genotype against Staphylococcus Aureus and E.coli infection and the predisposing role of the H8H21 genotype.4. Form the fluorescence quantitative real-time PCR (qPCR) analyses, the MBL1 gene was observed to be differentially expressed in bovine tissues (Fig.4). The liver of diseased cows had higher MBL1 expression than the ones with healthy (P＜0.05). However, no other significant associations were observed heart, spleen, lung, kidney, muscle and mammary tissue between the healthy samples and the diseased samples (P>0.05). It was low expression of MBL1 in spleen, lung, kidney, muscle and mammary and the less expression of MBL1 in spleen.