User:Andrew Yang/Notebook/MURI MIT/2012/02/06

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2/6/2012 To-Do

Summary of Weekend Progress

  • Restriction map confirmed that the restriction enzymes E, X, S all work
  • 4% gel confirmed the annealing of the ultramers for J85010, J85012, and J85013
  • Gel of the EX digests of J85201 and J06702; and the ES digests of S03119 and Chlor confirmed expected band lengths.
  • Gel extraction of the parts yielded good DNA concentrations, with good 260/230 ratio
  • Phosphatase->Gel extract->Ligate->Tranformed
  • Transformation results, however, yielded no colonies for all the negative controls and ligations. Postitive controls yielded conolonies
  • Because the ultramer annealing was confirmed, the ligation step must be the problem.
  • Will retry ligation following advice: http://openwetware.org/wiki/DNA_ligation

Today's Workflow

  • Miniprep J06702
  • PCR J06702 and 069 fragment with recently ordered Gibson primers J85014, J85015
  • Gibson assemble the PCR products
  • Reattempt ligation
  • Inoculate additional S03119.