A PCR plate is a multi-well plastic plate (usually 96, 384, or 48 wells) used to hold PCR reactions in a thermal cycler.
Follow these steps:
1. Preparation
Work in a clean area, preferably under a PCR hood. Label the plate and record a plate map. Thaw reagents on ice and mix gently.
2. Prepare the Master Mix
Calculate the total volume needed, plus about 10% extra. Mix buffer, dNTPs, primers, polymerase, and water. Add template DNA separately.
3. Dispense the Mix
Use a multichannel pipette. Add 15–25 µL master mix per well, then 1–5 µL template DNA. Change tips between samples to avoid contamination. Keep the plate on ice.
4. Seal the Plate
Use adhesive film, heat-sealing film, or cap strips. Press firmly to ensure a flat, tight seal with no wrinkles or gaps.
5. Spin Down
Centrifuge briefly (500–1000 rpm, 10–30 seconds) to collect liquid and remove bubbles.
6. Run the PCR
Place the plate in the thermal cycler, aligning the notch correctly. Set the program and start the run.
7. After the Run
Remove the plate carefully. For qPCR, read directly without removing the seal. Store at 4°C or –20°C if not analyzed immediately.
Key Precautions
- Wear gloves and use filter tips.
- Keep reagents on ice.
- Avoid bubbles.
- Seal tightly to prevent evaporation.
- Never reuse plates.
Common Plate Types
- 96-well: routine PCR
- 384-well: high-throughput
- White plates: qPCR
- Clear plates: standard PCR
Common Mistakes
Poor sealing, wrong plate type, fast pipetting, reusing plates, and not spinning down can all cause failed reactions.
If you share your application and thermal cycler model, more specific guidance can be provided.