Veterinary World

     Open access and peer reviewed journal  

ISSN (Online): 2231-0916

ISSN (Print): 0972-8988


Home l Editorial board l Instructions for authors l Reviewer guideline l Open access policy l Archives l FAQ

Open Access

Copyright: The authors. This article is an open access article licensed under the terms of the Creative Commons Attribution License

( which permits unrestricted use, distribution and reproduction in any medium, provided the work is properly cited.

Research (Published online: 04-12-2014)

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

A. Sakthivel Selvan, I. D. Gupta, A. Verma, M. V. Chaudhari and V. Kumar

Veterinary World, 7(12): 1037-1040



   doi: 10.14202/vetworld.2014.1037-1040



A. Sakthivel Selvan: Molecular Genetics Lab, Dairy Cattle Breeding Division, National Dairy Research Institute, Karnal - 132 001, Haryana, India;

I. D. Gupta: Molecular Genetics Lab, Dairy Cattle Breeding Division, National Dairy Research Institute, Karnal - 132 001, Haryana, India;

A. Verma: Molecular Genetics Lab, Dairy Cattle Breeding Division, National Dairy Research Institute, Karnal - 132 001, Haryana, India;

M. V. Chaudhari: Molecular Genetics Lab, Dairy Cattle Breeding Division, National Dairy Research Institute, Karnal - 132 001, Haryana, India;

V. Kumar: Molecular Genetics Lab, Dairy Cattle Breeding Division, National Dairy Research Institute, Karnal - 132 001, Haryana, India;


Received: 06-08-2014, Revised: 04-11-2014, Accepted: 07-11-2014, Published Online: 04-12-2014


Corresponding author: I. D. Gupta, e-mail:

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.

Keywords: cluster of differentiation 14, Hinf1, Karan Fries, mastitis, restriction fragment length polymorphism, single nucleotide polymorphism.

1. BAHS. (2012) Basic Animal Husbandry Statistics. Government of India, Ministry of Agriculture, Department of Animal Husbandry, Dairying and Fishries, Krishi Bhawan, New Delhi.
2. Gurnani, M., Sethi, R.K. and Nagarcenkar, R. (1986) Development of Karan Fries cattle at NDRI, Karnal. Dairy Information Bulletin No. 111.9. National Dairy Research Institute, Karnal, India: p1-2.
3. Shuster, D.E., Kehrli Jr, M.E., Ackermann, M.R. and Gilbert, R.O. (1992) Identification and prevalence of a genetic defect that causes leukocyte adhesion deficiency in Holstein cattle. Proc. Natl. Acad. Sci. USA, 89: 9225-9229.
PMid:1384046 PMCid:PMC50098
4. Qureshi, S., Larivičre, L., Sebastiani, G., Clermont, S., Skamene, E., Gros, P. and Malo, D. (1996) A high-resolution map in the chromosomal region surrounding the Lps locus. Genomics, 31: 283-294.
5. Maillard, J.C., Berthier, D., Chantal, I., Thevenon, S. and Sidibé, I. (2003) Selection assisted by BoLA-DR/DQ haplotype against susceptibility to bovine dermatophilosis. Genet. Sel. Evol, 35(1): 193-200.
6. Garcia-Briones, M.M., Russell, G.C., Oliver, R.A., Tami, C., Taboga, O., Carrillo, E., Palma, E.L., Sobrino, F. and Glass, E.J. (2000) Association of bovine DRB3 alleles with immune response to FMDV peptides and protection against viral challenge. Vaccine, 19: 1167-1171.
7. Takeda, H., Takami, M., Oguni, T., Tsuji, T., Yoneda, K., Sato, H., Ihara, N., Itoh, T., Kata, S.R. and Mishina, Y. (2002) Positional cloning of the gene LIMBIN responsible for bovine chondrodysplastic dwarfism. Proc. Natl. Acad. Sci., 99: 10549-10554.
PMid:12136126 PMCid:PMC124971
8. Kumar, N., Mitra, A., Ganguly, I., Singh, R., Deb, S.M., Srivatsava, S.K. and Sharma, A. (2005) Lack of association of brucellosis resistance with (GT)(13) microsatellite allele at 3'UTR of NRAMP1 gene in Indian zebu (Bos indicus) and crossbred (Bos indicus × Bos taurus) cattle. Vet. Microbiol., 111: 139-143.
9. Pinedo, P.J., Donovan, A., Rae, O. and DeLapaz, J. (2009) Association between paratuberculosis infection and general immune status in dairy cattle. Proceedings International Association for Paratuberculosis. College of Veterinary Medicine, Minneapolis, USA. p127.
10. Juliarena, M.A., Poli, M., Ceriani, C., Sala, L., Rodríguez, E., Gutierrez, S., Dolcini, G., Odeon, A. and Esteban, E.N. (2009) Antibody response against three widespread bovine viruses is not impaired in Holstein cattle bovine leukocyte antigen (BoLA) DRB3.2 carrying alleles associated with bovine leukemia virus resistance. J. Dairy. Sci., 92(1): 375-381.
11. Nascimento, C.S., Machado, A.M., Martinez, L.M., Silva, M.V.G., Martins, M.F. and Campos, A.L. (2006) Association of the bovine major histocompatibility complex (BoLA) BoLA-DRB3 gene with fat and protein production and somatic cell score in Brazilian Gyr dairy cattle (Bos indicus). Genet. Mol. Biol., 29(4): 641-647.
12. Rupp, R., Hernandez, A. and Mallard, B.A. (2007) Association of bovine leukocyte antigen (BoLA) DRB 3.2 with immune response, mastitis and production and type traits in Canadian Holsteins. J. Dairy. Sci., 90: 1029-1038.
13. Sugimoto, M., Fujikawa, A., Womack, J.E. and Sugimoto, Y. (2006) Evidence that bovine forebrain embryonic zinc finger-like gene influences immune response associated with mastitis resistance. Proc. Natl. Acad. Sci., 103(17): 6454-6459.
PMid:16611727 PMCid:PMC1458905
14. Wang, X., Xu, S., Xue, S., Ren, H. and Chen, J. (2007) Genetic polymorphism of TLR4 gene and correlation with mastitis in cattle. J. Genet. Genomics, 34(5): 406-412.
15. Wang, X., Xu, S., Xue, G., Li, J., Ren, H. and Luoren, Z. (2008) Cloning and SNP screening of the TLR4 gene and the association between its polymorphism and somatic cell score in dairy cattle. South. Afr. J. Anim. Sci, 38(2): 101-109.
16. Pant, S.D., Schenkel, F.S., Leyva-Baca, I., Sharma, B.S. and Karrow, N.A. (2007) Identification of single nucleotide polymorphisms in bovine CARD15 and their associations with health and production traits in Canadian Holsteins. BMC Genomics, 8: 421.
PMid:18005441 PMCid:PMC2234259
17. Soumya, N.P., Gupta, I.D., Verma, A., Raja, K.N., Chauhan, I. and Chaudhari, M.V. (2013) Identification of CARD-15 gene polymorphism in Sahiwal cattle. Indian J. Anim. Res., 47(1): 83-86.
18. Ibeagha-Awemu, E.M., Lee, J.W. and Ibeagha, A.E. (2008) Bovine CD14 gene characterization and relationship between polymorphisms and surface expression on monocytes and polymorphonuclear neutrophils. BMC Genet., 9: 50-60.
PMid:18691417 PMCid:PMC2536669
19. Beecher, C., Daly, M., Childs, S., Berry, D.P., Magee, DA., McCarthy, T.V. and, Giblin, L. (2010) Polymorphisms in bovine immune genes and their associations with somatic cell count and milk production in dairy cattle. BMC Genet., 11: 99-108.
PMid:21054834 PMCid:PMC3087511
20. Kumar, V., Gupta, I.D., Verma, A., Kumar, S.R. and Chaudhari M.V. (2014) CD14 gene polymorphism using HinfI restriction enzyme and its association with mastitis in Sahiwal cattle. Indian J. Anim. Res., 48: 11-13.
21. Ogorevc, J., Kunej, T., Razpet, A. and Dovč, P. (2009) Database of cattle candidate genes and genetic markers for milk production and mastitis. Anim. Genet., 40: 832-851.
PMid:19508288 PMCid:PMC2779988
22. Sambrook, J. and Russell, D.W. (1989) Preparation and analysis of eukaryotic DNA. In: Molecular Cloning: A Laboratory Manual. 3rd ed. Cold Spring Harbor Laboratory Press, New York. p6.1-6.62.