Uni-Directional Workflow
Definition
The physical principle governing forensic DNA laboratory layout: personnel and materials move from pre-PCR to post-PCR zones only, preventing high-copy-number amplified products from contaminating pre-amplification workspaces.
- Zones
- Pre-PCR and post-PCR, physically separated
- Movement rule
- Personnel and materials move pre-PCR to post-PCR only, never back
- Risk prevented
- Amplicon carryover contamination
- Field
- Forensic DNA laboratory design
Common questions
Why is amplicon carryover such a serious risk that it needs a whole workflow rule?+
PCR products exist in millions to billions of copies per reaction, so even a trace amount tracked back into a pre-amplification area on gloves, equipment or air currents can contaminate a fresh sample and produce a false profile.
What happens practically if someone needs to move from post-PCR back to pre-PCR?+
Standard practice requires a full change of lab coat, gloves and often physical exit and re-entry through a separate route, since the one-way design assumes no direct return path exists.
Related terms
- A-Addition
- The non-templated addition of a single adenine nucleotide to the 3' end of PCR products by Taq polymerase, exploited in forensic STR...
- AmpliTaq Gold
- A chemically modified hot-start form of Taq polymerase (Applied Biosystems) that is inactive at room temperature and requires a 10-11 minute activation...
- Annealing Temperature
- The reduced temperature (50-65°C) at which primers hybridise to single-stranded template. Determined by primer Tm; set to balance specificity against yield.
- Denaturation
- The separation of double-stranded DNA into two single strands by disrupting the hydrogen bonds between base pairs. In PCR, denaturation is achieved...
- Extension
- The PCR step at ~72°C at which thermostable DNA polymerase synthesises a new complementary strand from each primer, using dNTPs in the...
- Hot-Start Polymerase
- Any Taq formulation (antibody-mediated, chemically modified, or aptamer-blocked) that is inactive below ~60°C, preventing non-specific amplification during reaction setup.
- Melting Temperature (Tm)
- The temperature at which half of a primer-template duplex population is dissociated. Depends on primer length, GC content, and nearest-neighbour stacking interactions.
- Negative Amplification Control
- A no-template reaction well run alongside case samples; any amplified peak indicates reagent contamination or analyst-introduced contamination during setup.