No PCR Product (Complete Failure) (End-point PCR Primers)

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No bands visible on gel electrophoresis after PCR amplification Common causes: 1) NaCl at 50 mM inhibiting enzyme 2) Excessive KCl (>50 mM) in reaction 3) Incorrect annealing temperature 4) Incomplete denaturation (insufficient time or temperature) 5) GC-rich template regions requiring potassium-free conditions 6) 10% DMSO partially inhibiting Taq 7) Hemin in blood samples inhibiting Taq 8) Super-irradiated mineral oil (>2500 mJ/cm2) 9) Use of wooden toothpick for sample transfer 10) Presence of Taq inhibitors (indigo dyes, heme) Solutions: 1) Ensure NaCl does not exceed 50 mM 2) Keep KCl concentration below 50 mM 3) Optimize annealing temperature 4) Increase denaturation time and ensure temperature is sufficiently high 5) For GC-rich templates, eliminate potassium from reaction 6) Reduce DMSO concentration below 10% 7) Dilute blood samples or use purified DNA to reduce hemin 8) Use mineral oil treated with ≤2500 mJ/cm2 9) Avoid wooden toothpicks; use sterile plastic tips instead 10) Add BSA, increase Taq amount, or increase reaction volume to dilute inhibitors