Rapid and Sensitive Detection of BLAD in Cattle Population
Keywords:
BLAD, CD18, genotyping, high-resolution meltingAbstract
Bovine leukocyte adhesion deficiency (BLAD) is an autosomal recessive disorder with negative impact on dairy cattle breeding. The molecular basis of BLAD is a single point mutation (A→G), resulting in a single amino acid substitution (aspartic acid → glycine) at amino acid 128 in the adhesion molecule CD18. The object of this study was to establish a fast and sensitive molecular genotyping assay to detect BLAD carriers using high-resolution melting (HRM) curve analysis. We tested animals with known genotypes for BLAD that were previously confirmed by PCR-RFLP method, and then examined the sensitivity of mutation detection using PCR followed by HRM curve analysis. BLAD carriers were readily detectable using HRM assay. Thus, the PCR-HRM genotyping method is a rapid, easily interpretable, reliable and cost-effective assay for BLAD mutant allele detection. This assay can be useful in cattle genotyping and genetic selection.
References
Odalys Uffo, Atzel Acosta, Molecular diagnosis and control of genetic bovine diseases, Biotecnologia Aplicada, 2009, 26 (3), 204-208
Shuster D.E., Kehrli M.E, Ackermann M.R., Gilbert R.O., Identification and prevalence of a genetic defect that causes leukocyte adhesion deficiency in Holstein cattle, Proc. Nati. Acad. Sci. USA, 1992, 89 (19), 9225-9229
Citek J., Rehout V., Hajkova J., Pavkova J., Monitoring of the genetic health of cattle in the Czech Republic, Vet. Med., 2006, 51(6), 333-39
Radi Z.A, Kehrli M.E, Ackermann M.R., Cell adhesion molecules, leukocyte trafficking, and strategies to reduce leukocyte infiltration, J. Vet. Intern. Med., 2001, 15, 516–529
Kishimoto K.K., Hollander N., Roberts T.M., Anderson D.C., Springer T.A., Heterogeneous mutations in the β subunit common to the LFA-1, Mac-1, and p150,95 glycoproteins cause leukocyte adhesion deficiency, Cell, 1987, 50, 193-202
Hagemoser W.A., Roth J.A., Löfsted J., Fagerland J.A. Granulocytopathy in a Holstein heifer. J. Am. Vet. Med. Assoc., 1983, 183,1093-4
Nagahata H., Bovine leukocyte adhesion deficiency (BLAD): A Review. Journal of Veterinary Medical Science, 2004, 66(12), 1475–1482
Ribeiro L.A., Baron E.E., Martinez M.L., Coutinho L.L., PCR screening and allele frequency estimation of bovine leukocyte adhesion deficiency in Holstein and Gir cattle in Brazil, Genet. Mol. Biol., 2000, 23 (4), doi: 10.1590/S1415-47572000000400021
Rahimi G., Nejati-Javaremi A., Olek K., Genotyping BLAD, DUMPS and κ-CSN Loci in Holstein Young Bulls of the National Animal Breeding Center of Iran, Pakistan Journal of Biological Sciences, 2006, 9 (7), 1389-1392
Vatasescu-Balcan R.A., Manea M.A., Georgescu S.E., Dinischiotu A., Tesio C.D., Costache M., Evidence of single point mutation inducing BLAD disease in Romanian Holstein-derived cattle breed, Biotechnology in Animal Husbandry, 2010, 23(5-6), 375-382
Akyuz B., Okan E., Ozgecan K.A., Detection of Bovine Leukocyte Adhesion Deficiency (BLAD) Allele in Holstein Cows Reared in Kayseri Vicinity, Kafkas Univ Vet Fak Derg, 2010, 16(2), 519-521, http://vetdergi.kafkas.edu.tr/extdocs/2010_3/519_521.pdf
Meydan H., Yildiz M.A., Agerholm J.S., Screening for bovine leukocyte adhesion deficiency, deficiency of uridine monophosphate synthase, complex vertebral malformation, bovine citrullinaemia, and factor XI deficiency in Holstein cows reared in Turkey, Acta Vet. Scand, 2010, 52(1), 56, doi: 10.1186/1751-0147-52-56
Ilie D.E., Gavriliuc O.I., Cean A., Neamț R.I., Neciu F.C., Păcală N, Control strategies for prevention of undesirable traits in cattle, Scientific Papers: Animal Sciences and Biotechnologies, 2011, 44(1), 415-419
Reed G.H., Wittwer C.T., Sensitivity and specificity of single-nucleotide polymorphism scanning by high-resolution melting analysis, Clin. Chem., 2004, 50, 1748-1754
Montgomery J., Wittwer C.T., Palais R., Zhou L., Simultaneous mutation scanning and genotyping by high-resolution DNA melting analysis, Nat Protoc, 2007, 2(1), 59–66
Zsolnai A., Fesus L., Enhancement of PCR- RFLP Typing of Bovine Leukocyte Adhesion Deficiency, BioTechniques, 1997, 23,380-382
Balic M., Pichler M., Strutz J., Heitzer E., Ausch C., Samonigg H., Cote R.J., Dandachi N., High quality assessment of DNA methylation in archival tissues from colorectal cancer patients using quantitative high-resolution melting analysis, J Mol Diagn., 2009, 11(2), 102-8
Ilie D.E., Cean A., Gavriliuc O.I., Carstea A.C., Grădinaru A.C., High-Resolution Melting Assay as a Tool for Identification of CSN3 Genotypes in Cattle Population, Scientific Papers: Animal Sciences and Biotechnologies, 2013, 46 (1), 102-105
Santos S., Marques V., Pires M., Silveira L, Oliveira H., Lança V., Brito D., Madeira H, Esteves J.F., Freitas A., Carreira I.M., Gaspar I.M., Monteiro C., Fernandes A.R., High resolution melting: improvements in the genetic diagnosis of hypertrophic cardiomyopathy in a Portuguese cohort, BMC Medical Genetics 2012, 13, 17
