Complete qRT-PCR Workflow Protocol Step-by-Step

critical

Complete Absence of PCR Product

No bands visible on agarose gel after PCR amplification, indicating complete reaction failure.

💡 6 · ✓ 6

severe

No or Low PCR Amplification Due to Plastic Consumable Issues

PCR reaction fails to produce expected amplification or yields significantly reduced product, despite optimized reagents and cycling parameters. The issue traces to suboptimal thermal transfer or contamination from the plastic vessel itself.

💡 4 · ✓ 4

severe

PCR Sample Evaporation and Loss During Cycling

Visible reduction in reaction volume after thermal cycling, with condensation on tube caps or film. May result in concentrated reagents, failed reactions, or inability to recover product.

💡 3 · ✓ 3