Team:Cooper Union/Protocols

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Cooper Union 2014 iGEM




Common Laboratory Protocols









Bacterial Transformation






Colony PCR






Gel Extraction






Miniprep






PCR Purification






PCR Reaction






Restriction Enzyme Digest



Materials
10x Buffer (Compatible to the enzyme)
Enzyme
BSA - diluted to 10x
ddH2O
DNA - up to 1μg per reaction.

Procedure Verify all times and temperatures for specific enzyme prior to use. This is general protocol.
  1. Prewarm 2 water baths to 37°C and 70°C
  2. Thaw buffers, diluted BSA. Keep all reagents on ice
  3. Set up reactions based on table below.
    ReagentVolumeExample
    Reaction Buffer1/10 total volume2μL
    Diluted BSA (10x)1/10 total volume2μL
    DNAcalculate for target ng3.7μL
    ddH2Ovolume up to (total volume - 1μL)11.3μL
    Enzymegenerally 1μL1μL
    Total Volume20uL
  4. Incubate 60min 37°C.
  5. Heat inactive 70°C for 15 min.
  6. Store in -20°C or use immediately.


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TdT Heat Inactivation






TdT Nucleotide Addition






Yeast Gene Knockout






Yeast Mating






Yeast Sporulation/Zymolyase Treatment