Designing multiplex PCR tests for simultaneous screening of bovine leukocyte adhesion deficiency

Title: Designing multiplex PCR tests for simultaneous screening of bovine leukocyte adhesion deficiency, bovine citrullinemia and factor XI deficiency genetic diseases in cattle

Authors: Anshuman Kumar, Ravi Kumar D, Vineeth MR, Govind Mohan, S Jayakumar, Saket K Niranjan and ID Gupta

Source: Ruminant Science (2017)-6(2):215-220.

Cite this reference as: Kumar Anshuman, Kumar D Ravi, MR Vineeth, Mohan Govind, Jayakumar S, Niranjan SK and Gupta ID (2017). Designing multiplex PCR tests for simultaneous screening of bovine leukocyte adhesion deficiency, bovine citrullinemia and factor XI deficiency genetic diseases in cattle. Ruminant Science 6(2):215-220.


Bovine Leukocyte Adhesion Deficiency (BLAD), Bovine Citrullinemia (BC) and Factor XI deficiency (FXID) are most prevalent lethal genetic disorders in cattle, causing death of homozygote animal at young stage and severe adverse effects on viability in heterozygous condition also. These disorders propagate unabated in absence of screening of carrier bulls in herds, worldwide. After identifying the loci for these disorders, the breeding animals; especially bulls are being screened separately through DNA based tests in various countries including India. Under this study, an attempt was made to detect the BLAD, BC and FXID mutations simultaneously after multiplexing of these conventional PCR based DNA tests. Two multiplexed PCR protocols – one for BLAD and FXID and another for BC and FXID loci screening were developed. Along with modifying the primers, the PCR components and conditions were also standardized for developing both multiplexed PCR protocols. In both of the protocols, loci were amplified and further analysed by restriction fragment length polymorphism (RFLP) using TaqI and AvaII restriction enzymes, respectively. After PCR-RFLP, the products were resolved in gel electrophoresis, which were easily identifiable to ascertain their genotypes.  These protocols produced the results without compromising accuracy, reducing overall time and cost incurred in separate PCR tests and may be used for rapid screening of these genetic disorders in a herd.


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