Rui:LabNotes/Haplotyping/2011-8-3

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Methylation haplotyping[edit]

  • Repeat the protocol used in lab note on 7/13/11

Cells[edit]

  • Cells from Alan: GM12878_P18
  • Cell counting machine: Dr. Varghese lab (3rd floor)
  • 100ul cell culture mixed with 10ml counting buffer, put on holder and start
  • 152,116 cells/ml in 1XPBS
  1. Mix well, take 1ml cells to spin down 800g (3k rpm) 3min --> no pellet
  2. Additional 3min --> no pellet
  3. take 2ml cells to spin down 5k rpm 5min --> no pellet
  4. Unsure about the cell condition, aliquot anyway and will test few tubes (will ask Alan for more cells)

Solutions[edit]

  • Feel confused about Alice's lysis buffer, decide to remake 10X NP40 lysis buffer
  • Qiagen protease has contamination, reorder, resuspend (5Au/ml) and aliquot for everyone
  1. Make 1M MgCl2 solution, filter with 0.22um filter
  2. Make 10ml 10X NP40 lysis buffer [100mM Tris.Hcl pH 7.5, 100mM NaCl, 30mM MgCl2, 1% NP40, Crawford et al. PNAS 2004]
  3. Make 1ml 1X lysis buffer + 40ul protease (0.2Au/ml final)