1Sandor Animal Biogenics Pvt. Ltd, Banjara Hills, Hyderabad, India
2Sandor Life Science Pvt. Ltd, Banjara Hills, Hyderabad, India
Receive Date: 06 April 2014,
Revise Date: 20 May 2014,
Accept Date: 31 May 2014
In India, Holstein and its crosses are being used extensively in breeding programmes and all these breeding bulls are screened for autosomal recessive genes. Blood samples are collected in ethylenediaminetetraacetic acid (EDTA) coated tubes and DNA was isolated by using phenol-chloroform method. Polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) wereperformed by using bovine leukocyte adhesion deficiency (BLAD) specific primersand TaqI restriction enzyme for diagnosis of bovine leukocyte adhesion deficiency (BLAD) syndrome. The bull was found to be heterozygous for BLAD allele. The PCR product was sequenced by automated sequencer using the ABI big dye Ver 3.1 for detection of mutation at position 383 bp (A/G). Sequence analysis comparison was performed using the codon code aligner 4.0.4 software. The sequence of carrier animal confirmed polymorphism at 383 bp position. The sequence was also compared with sequence of normal Holstein as a control and the sequence available with NCBI (accession No. NC-007299). The comparison of sequences revealed a heterozygous polymorphism at 348 position (T>C) in a carrier animal whereas in homozygous in a control Holstein which was normal for BLAD. The new polymorphism at 348 position was found to be silent as it does not change amino acid (asparagine, AAT>AAC) within exon 4 of CD18 gene. The partial sequence of new polymorphism / silent mutation has already been submitted to NCBI (accession No. KF840683). Further studies have to be carried out to elucidate the possible association of the CD18 silent point mutation at 348 bp position as a potential molecular marker for milk production traits.
Andrews A.H., Fishwick J. and Waters R.J. (1996). Bovine leukocyte adhesion deficiency (blad) in a one year old Holstein Friesian bull-the first report in the United Kingdom. Br. Vet. J. 152, 347-351. Bell J.R. (2008). A simple way to treat PCR products prior to sequencing using ExoSAP-IT. Bio. Tech. 44, 834-834. Citek J., Rehout V., Hajkova J. And Pavkova J. (2006). Monitoring of the genetic health of cattle in the Czech Republic. Vet. Med. 51, 333-339. Czarnik U. (2000). Breeding and genetic population aspects of the occurrence of bovine leukocyte adhesion deficiency (BLAD) in black-and-white cattle. Dissert. Monogr. 32, 1-44. CzarnikU. And Kaminski S. (1997). Detection of intronic sequennce in the bovine ITGB2 gene. Anim. Genet. 28, 320-321. Czarnik U., Galinski M., Pareek C.S., Zabolewicz T. and Wielgosz-Groth Z. (2007). Study of an association between SNP 775 C>T within the bovine ITBG2 gene and milk performance traits in Black and white cows. Czech J. Anim. Sci. 52, 1-6. Huang Y.C., Chang H.L., Lin D.Y., Liaw R.B. and Chen J.C. (2000). Gene frequencies of bovine leukocyte adhesion deficiency of Holsteins in Taiwan. J. Taiwan Livest. Res. 33, 37-47. Hynes R.O. (1987). Integrins: a family of cell surface receptors. Cell. 48, 549-554. Janosa A., Dohy J. and Baranyai B. (1999). Comparison of milk production of the progeny of BLAD-carrier and healthy Holstein bulls in Hungary. Acta Vet. Hung. 47, 283-289. Kriegesmann B., Jansen S., Baumgartner B.G. and Brenig B. (1997). Partial genomic structure of the bovine CD18 gene and the refinement of test for bovine leukocyte adhesion deficiency. J. Dairy Sci. 80, 2547-2549. Larson R.S. and Springer T.A. (1990). Structure and function of leukocyte integrins. Immunol. Rev. 114, 181-217. Li J., Wang H., Zhang Y.,Hou M.,Zhong J. and Zhang Y.(2011). Identification of BLAD and citrullinemia carriers in Chinese Holstein cattle. Anim. Sci. Pap. Rep. 2, 37-42. Malech H.L. and Gallin J.I. (1987). Neutrophils in human diseases. New Engl. J. Med. 317, 687-694. Nagahata H., Miura T., Tagaki K., Ohtake M., Noda H., Yasuda T. and Nioka K. (1997). Prevalence and allele frequency estimation of bovine leukocyte adhesion deficiency (BLAD) in Holstein-Friesian cattle in Japan. J. Vet. Med. Sci. 59, 233-238. Nagahata H., Noda H., Takahashi K., Kurosawa T. and Sonoda M. (1987). Bovine granulocytopathy syndrome: neutrophil dysfunction in Holstein Friesian calves. J. Vet. Med. Ser A.34, 445-451. Norouzy A., Nassiry M.R., Shahrody F.E., Javadmanesh A., Abadi M.R.M. and Sulimova G.E. (2005). Identification of bovine leukocyte adhesion deficiency (BLAD) carriers in Holstein and Brown Swiss AI bulls in Iran. Genetica. 41, 1697-1701. Padeeri M., Vijaykumar K., Grupe S., Narayan M.P., Schwerin M. and Kumar M.H. (1999). Incidence of hereditary citrullinemia and bovine ieucocyte adhesion deficiency syndrome in Indian dairy cattle (Bos taurus and Bos indicus) and buffalo (Bubalus bubalis) population. Arch. Tierz. Dummerstorf. 42, 347-352. Patel M., Patel R.K., Singh K.M., Rank D.N., Thakur M.C. and Khan A. (2011). Detection of genetic polymorphism in CD18 gene in cattle by PCR-RFLP. Wayamba J. Anim. Sci. 3, 110-111. Patel R.K., Singh K.M., Soni K.J., Chauhan J.B. and Rao K.S.R. (2007). Low incidence of bovine leukocyte adhesion deficiency (BLAD) carriers in Indian cattle and buffalo breeds. J. Appl. Genet. 48, 153-155. Ribeiro L.A., Baron E.E., Martinez M.L. and Coutinho L.L. (2000). PCR screening and allele frequency estimation of bovine leukocyte adhesion deficiency in Holstein and Gir cattle in Brazil.Genet. Mol. Biol. 23, 831-834. Roy A., Kotikalapudi R., Patel R.K., Anantaneni R. And Katragadda S. (2012). New cases of bovine leukocyte adhesion deficiency (BLAD) carriers in Indian Holstein cattle. Int. J. Vet. Sci. 1, 80-82. Sambrook J., Fritsch E.F. and Maniatis T. (1989). Molecular Cloning:A Laboratory Manual. Cold Spring Harbor Laboratory Press. New York, USA. Shuster D.E., Kehrli 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. 89, 9225-9229. Taralik K. (1998). Association between BLAD carrier and non BLAD carrier of Holstein-Friesian bulls and the production of their daughters. Arch. Fuer. Tierz. 41, 339-344. Yathish H.M., Sharma A., Kumar V., Jain A. and Chakraborty D. (2010). Genetic polymorphism of CD18 gene in Karan Fries young bull calves using PCR-RFLP analysis. Curr. Trend. Biotechnol. Pharm. 4, 900-907.