Molecular identification and characterization of Muscid flies (Diptera: Muscidae) of medico-veterinary importance from the Gangetic plains of Eastern India

DNA Barcoding of Economically Important Fruit Flies (Diptera: Tephritidae) from the Lower Gangetic Plains of Eastern India

“…Amplification of about 700 base pairs from the 5' end of the mitochondrial COI gene was performed using roughly 20 ng of genomic DNA employing primers forward LCO-1490 (F) (GGT CAA CAA ATC ATA AAG ATA TTG G) and reverse HCO-2198 (R) (TAA ACT TCA GGG TGA CCA AAA AAT CA) [34]. PCR was performed in a 50µl total reaction volume comprising 20 Pico moles of each primer [34a], 100 mM KCl, 20 mM Tris-HCl (pH 8.0), 1 mM DTT, 0.1 mM EDTA, 2.0 mM MgCl2, 0.25 mM of each dNTP, primer cocktail, and 1U of Taq polymerase (Takara BIO Inc., Japan) with the following cycling parameters: 5 min at 94•C; followed by 40 cycles of 30 s at 94•C, 40 s at 53•C, 1 min at 72•C and final extension for 5 min at 72•C [33,35]. To verify the amplicon's size, the amplified products were observed on a 1% agarose gel, dyed with SYBR@safe DNA gel dye, and captured on an Invitrogen-safe gel imager.…”