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During PCR Reagent preparation, if DNA fragments from the lab environment, such as a DNA template amplified in a previous qPCR experiment, enter the qPCR reaction or reagents (even in small quantities), they can be amplified during the reaction. This contamination and non-specific amplification can cause misleading results, such as false positives. Hence, contamination needs to be rigorously controlled.

 

Do’s and Don’ts for Contamination Control during PCR Reagent Preparation 

 

Do’s

  • Change shoes or use shoe covers, before entering the Reagent Preparation/ Master Mix Room.
  • Always clean the work area properly before and after completing the work, using disinfectants.
  • Keep master mix reagents in -20°C freezer.
  • Label tools and instruments to be used only in Master Mix Room. ​
  • Use 0.5 ml individual PCR tubes.​
  • Use a PCR hood.
  • Decontaminate and calibrate pipettes regularly.​
  • Always prepare fresh disinfectant (1% sodium hypochlorite solution​).
  • Establish regular (e.g., weekly) and thorough laboratory cleaning protocols for floors, doors, walls.

 

Don’ts

  • Tools and instruments used in Master Mix Room should not be used elsewhere.
  • After working in other areas, do not come back and clean the reagent preparation area.

 

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