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Protocol/17
From 2010.igem.org
Protocol 17: PCR
Before you start:
- KEEP EVERYTHING ON ICE. Put Taq back into freezer as soon as you`re done with it. DON`T put back dNTP tubes.
- Reserve a thermocycler and check what size of tubes it takes.
- Making a master mix conserves expensive reagents, so try to always use one.
- You may have to do dilutions of your reagents in order to make them usable for PCR (ex: primers, plasmid DNA...)
- DON'T PUT PRIMERS OR TEMPLATE INTO THE MASTER MIX. ADD POLYMERASE TO THE MASTER MIX LAST (after your other tubes already have template DNA and primers in them)!
Protocol:
- PCR buffer 5ul
- 10uM dNTP's 1ul
- 50uM MgCl2 2ul
- forward primer 2.5ul
- reverse primer 2.5ul
- 1ng template DNA 1ul
- Taq polymerase 0.5ul
- MilliQ (H2O) 35.5ul (to make total volume 50ul)
TOTAL 50ul
