Cluster of differentiation 14 gene polymorphism and its association with incidence of clinical mastitis in Karan fries cattle

Aim: The present study was undertaken with the objectives to characterize, identify DNA polymorphism in cluster of differentiation 14 (CD14) gene in Karan Fries (KF) cattle and to analyze association between genetic variants with incidence of clinical mastitis in National Dairy Research Institute (NDRI) herd, Karnal. Materials and Methods: Genomic DNA was extracted using blood of randomly selected hundred KF lactating cattle by phenol-chloroform method. After checking its quality and quantity, polymerase chain reaction (PCR) was carried out using reported primers to amplify 832 base pair region covering nucleotide base position number 1012 to 1843 (part of promoter, 5’UTR, exon 1, intron 1 and part of exon 2) of bovine CD14 gene. The PCR amplified target product was purified, sequenced and further ClustalW analysis was done to align edited sequence with reported Bos taurus sequence (EU148610.1). The restriction fragment length polymorphism (RFLP) analysis was performed for each KF cow using HinfI restriction enzyme (RE). Cows were assigned genotypes obtained by PCR-RFLP analysis and association study was done using Chi-square (χ2) test. Results: After PCR amplification, DNA sequencing of amplicon confirmed the 832 bases covering 1012 to 1843 nucleotide base position of bovine CD14 gene. ClustalW multiple sequence alignment program for DNA revealed six nucleotide changes in KF cows at positions T1117D, T1239G, T1291C, G1359C, G1361A, and G1811A. Cows were also screened using PCR-RFLP with HinfI RE, which revealed three genotypes CC, CD and DD that differed significantly regarding mastitis incidence. Within CC genotype, 72.73% of cows were in a mastitis non-affected group whereas, those in CD and DD genotypes 69.44% and 60.38% respectively were mastitis affected. Conclusion: KF cows with allele C of CD14 gene were less susceptibility to mastitis compared with D allele.