Species-Specific PCR Assay for Identification of Buffalo Meat

Two primer sets based on mitochondrial D-loop and cytochrome b gene sequences of buffalo with amplicon size of 227 and 152 bp, respectively, were opted for species identification of buffalo meat. The buffalo specific primer was subsequently tested for cross reactivity with cattle (ox), sheep, goat and pig meat. Primer targeting D-loop gene, yielded amplicon with buffalo meat as well as yielded size fragment with sheep and goat meat. However, with the primer based on cytochrome b gene, an intense band in buffalo meat and, a relatively faint band of similar size fragment in sheep was observed. In remaining three species cattle, goat and pig, no amplification was observed at usual 600C annealing temperature. In order to eliminate the cross reactivity of buffalo specific primer in sheep, amplification was attempted at the higher annealing temperatures 64.4oC and 67oC. At the annealing temperatures of 67oC, no cross reactivity of the buffalo specific primer was obtained in sheep as well as cattle (ox), goat and pig meat and only species-specific fragment was obtained in buffalo meat. Thus, buffalo species specific primer based on cytochrome b gene, under modified PCR condition was found useful.