• Created by Unknown User (asjm@mit.edu), last modified by Unknown User (krbrink@mit.edu) on Jun 27, 2014 09:06
To do:
  • High school presentations
  • Re-culture Syk and Lyn
  • Update our section of the parts list
  • Design Lyn-TEVp fusion

Notes:

  • Syk and Lyn concentrations found to be only ~15 ng/uL, so need to grow new cultures and re-miniprep
  • Primers iGEM2014-EE-[003,006] found to be missing spacer before BsaI site; corrected primers re-ordered
To do:
  • Miniprep Syk/Lyn and nanodrop
  • Sequence Syk/Lyn
  • Discuss antibody orders with Brian
  • NEGEM
To do:
  • All the PCRs!
    • CD79A + stop codon
    • CD79A + TCS
    • mKate (for fusion proteins)
  • Sequence Syk/Lyn
To do:
  • Image gel from June 22
  • Golden Gate PCRs from June 22
  • More PCRs with new primers

To do:

  • Redo Syk-eYFP PCR with DMSO
  • Image gel for yesterday's PCRs
  • Purify and gel Syk-eYFP PCR
  • Run remaining PCRs
  • Purify and gel remaining PCRs
  • Golden Gates
  • Put gel images on parts pages
  • Update wiki

Notes:

  • All of the original primers have arrived.
  • CD79B PCRs failed again, this time with DMSO in the mix.
  • New CD79B primers are designed and ordered.
  • When loading the gel, the Lyn wouldn't fill the wells, but instead seemed to float on top of the TAE. The reason for this is currently unknown, but it probably needs to be redone.

 

To do:

  • Redo Syk-15aa and Lyn PCRs with DMSO
  • Redo CD79B PCRs with GC buffer and original primers
  • Purify and gel PCR products
  • Run DpnI digest on TEVp product
  • Golden Gates

Notes:

To do:
  • Grow up colonies from Golden Gates of CD79A, CD79A-TCS-G4, Syk+stop
    • Sequence these (need sequencing primers for CD79A-TCS-G4)
  • Transform Golden Gates of Lyn+stop, Lyn-mKate, Lyn-TEVp, Syk-TEVp, Syk-eYFP
  • PCR Gmab Heavy and Light chains
  • Design sequencing primers for everything
  • Order antibodies
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