Team:Baltimore/Notebook

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Notebook

Run through for abstract and scheduling:

Start with plasmid with taq gene and pst1 gene

Remove pst1 site from taq coding sequence

  • In order to remove the restriction site, do site directed mutagenesis (1day)
  • PCR
  • Digestion
  • Transformation
  • Screening (1day)
    • Dilute DNA
    • Colony PCR individually
    • Digestion of PCR product with pst1
    • Run gel ~2500bp

Add biobrick prefix/suffix to taq coding sequence

  • PCR and add the prefix and suffix as primers (one prefix has an extra AG-make sure to use the correct one) (1-2 days)
    • Run gel
    • Cut out of gel
  • Cut with restriction enzyme
  • Clean up DNA
  • Ligation with vector (could be vector with the terminator sequence, promoter and RBS)
  • Transformation

Add a promoter, transcriptional terminator, ribosome binding site (RBS)

  • Screen colonies (1 day)
    • Colony PCR
    • Restriction Digestion
    • Clean up DNA
  • Sequence (1 day)

Make taq protein

Compare it to other enzymes and make sure it works