User:Karmella Haynes/Notebook/PcTF Genomics/2015/06/25

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mm/dd/yy

  • Cas-tone - cell transfections

Cas-tone - transfections

  • Final H3#_MV11 builds verified on 6/24/15
  • Co-transfect H3#_MV11 with pSP-guide RNA plasmid (built by Ben Nyer)
  • Use Luc14 cells (from Rene)
  1. H340_MV11 + pSP###
  2. H360_MV11 + pSP###
  3. H380_MV11 + pSP###
  4. H3116_MV11 + pSP###
  5. H3136_MV11 + pSP###
  6. pSP### (negative)
Well H3#_MV11 (1.0 μg) pSP# (1.0 μg) OptiMEM PLUS rgt. Lipo LTX
1 # # 570.0 2.5 7.5
2 # # 570.0 2.5 7.5
3 # # 570.0 2.5 7.5
4 # # 570.0 2.5 7.5
5 # # 570.0 2.5 7.5
6 # # 570.0 2.5 7.5
  • Pre-warm ps-free DMEM
  • Make 20 μL plasmid/dH2O mixes in sterile 1.5 ml tubes
  • Add OptiMEM
  • Add PLUS --> 5 min./ RT
  • Add Lipo LTX --> 30 min./ RT
  • Refresh growth medium w/ ps-free DMEM
  • Add 600 μL complexes
  • Incubate overnight



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