FAQ: Is Monarch-purified gDNA suitable for long-range PCR?

Yes. The large fragment size of Monarch-purified gDNA makes it excellent starting material for long range PCR approaches. For long range PCR, we recommend LongAmp TaqDNA Polymerase (NEB #M0323), LongAmp Taq 2x Master Mix (NEB #M0287) or LongAmp Hot Start Taq Polymerase (NEB #M0534).

A. Amplification reactions were set up with primer pairs specific for 6, 8, 10, 12, 16, 20 kb amplicons from human DNA. LongAmp® Hot Start Taq 2x Master Mix (NEB #M0533) was used and 25 ng template DNA was added to each sample. PCR reactions were carried out on an Applied Biosystems 2720 Thermal Cycler. Monarch-purified genomic DNA isolated from HeLa cells and human blood were compared to commercially available reference DNA from the human cell line NA19240 F11. 10 of 20 μl was loaded on a 1.5% agarose gel, using the 1 kb DNA Ladder (NEB #N3232) as a marker. Results indicated DNA was of high-integrity and suitable for long range PCR.

B. Monarch-purified genomic DNA from human whole blood, HeLa cells and mouse tail was diluted to produce a five log range of input template concentrations. The results were generated using primers targeting gHEME (human whole blood) and gREL (HeLa, mouse tail) for qPCR assays with the Luna Universal qPCR Master Mix (NEB #M3003) and cycled on a Bio-Rad® CFX Touch qPCR thermal cycler. Results indicated that DNA is highly pure and free from inhibitors, optimal for qPCR.