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De novo oligonucleotide primers for improved detection of a diagnostic single nucleotide polymorphism within CD18 sub unit of beta 2 integrin gene in cattle

BLAD is an economically important genetic disorder in cattle. The incidence rate in India is currently reported to be 3.23%. This study was therefore aimed to design a new set of improved PCR primers that can be used for convenient detection of the diagnostic SNP at codon 128 of the CD18 gene of cattle responsible for causing the disease, using Taq I enzyme-mediated RFLP analysis. In depth, bioinformatic analysis was performed to confirm the suitability of primers to substitute published versions that generated shorter fragments and required polyacrylamide gel for detection. The new set of primers generated 184 bp am- plicon with an asymmetrically placed diagnostic Taq I recognition site that cleaved it into 132 bp and 52 bp, respectively. It was demonstrated that the 132 bp and 184 bp fragments were sufficient to diagnose carriers excluding the need for identifying the 52 bp fragment in a regular, 3% agarose gel run on 0.5% TBE instead of conventional non-denaturing polyacrylamide gel.

Author(s): PN Mukhopadhyaya

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