FAQ: When using LunaScript cDNA products in downstream PCR (i.e., two-step RT-PCR), what volumes and PCR products can be used? 

The cDNA products of LunaScript reactions are compatible with many PCR reagents. Typically, we recommend OneTaq 2X Master Mix (NEB #M0482 or NEB #M0485) for PCR detection up to 5kb, Q5 Hot Start High-Fidelity 2X Master Mix (NEB #M0494) for highest fidelity, and LongAmp Taq 2X Master Mix (NEB #M0287) for high yields from longer products.

Depending on the targets/primers, the optimal cDNA input may vary and should be optimized but typically, we recommend 1 μl of the cDNA reaction as input into a 25 μl PCR. Where useful, up to 20% PCR volume can be added (e.g. 5 μl cDNA product in a 25 μl PCR).