Agarose Gel Electrophoresis of DNA Amplicons Post PCR: A Method to Analyze Products of Multiplex PCR and Evaluate PCR Reaction Success
Transcript
According to the manufacturer's recommendations, add agarose gel powder in 1x TBE buffer to reach 1.5 (weight/volume) percent, and heat to dissolve.
Prior to casting, tilt the dissolved gel solution briefly under running water. Add 5 microliters of fluorescent nucleic acid dye per 50 microliters of gel solution, and mix.
Assemble and level the casting tray. Add combs as appropriate for the number of samples. Pour the gel solution into the cast, and wait for it to solidify.
After that, submerge the cast containing the gel in 1x TBE buffer in a gel electrophoresis system. Mix 5 microliters of the PCR products together with 2 microliters of loading dye in new PCR strips. Transfer 5 microliters of the mixtures to the wells in the gel. Save at least one well empty. Add 5 microliters of DNA ladder in the empty well for reference.
Plug in the electrodes and run the gel at 110 volts per 15 centimeters for approximately 1 hour. Use a UV-based gel imaging system to verify the presence of multiple bands representing PCR amplicons.
