Jie:LabNotes/CpgSeq/2009-5-20

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bisulfite conversion of PGP samples

sample sample concentration sample volumn ddH2O conversion reagents conversed DNA concentration and volumn
09_1 NA21687*1 062507 241.1ng/ul x 1 tubes 8ul 12ul 130ul 95.4ng/ul x 10ul
09_2 NA21781*1 102307 240ng/ul x 1 tubes 8ul 12ul 130ul 120.2ng/ul x 10ul
09_3 NA21833*1 101807 253.9ng/ul x 1 tube 8ul 12ul 130ul 121.5ng/ul x 10ul
09_4 NA21660*1 053007 243.2ng/ul x 1 tube 8ul 12ul 130ul 120.3ng/ul x 10ul
09_5 NA21731*1 101807 217.4ng/ul x 1 tube 10ul 10ul 130ul 127.3ng/ul x 10ul
09_6 NA21846*1 100407 186.5ng/ul x 1 tube 11ul 9ul 130ul 151.3ng/ul x 10ul
09_7 NA21070*1 100406 329.7ng/ul x 1 tube 6ul 14ul 130ul 311.2ng/ul x 10ul
Jurkat gDNA 140ng/ul x 3 tube 14ul 6ul 130ul 255.6ng/ul x 20ul


  • Add 900ul ddH2O, 300ul M-Dilution Buffer and 50ul M-Dissolving Buffer to a tube of CT Conversion Reagent.
  • Add 130ul of CT conversion reagent to samples.
  • 98C 10min -> 64C 150min -> 4c overnight.
  • Add 600ul M-Binding buffer to spin columns.
  • Add the converted samples to the columns, close the gap and mix by inverting several times.
  • Spin at 15,000rpm for 30sec.
  • Add 100ul M-Wash Buffer, spin for 30 sec, discard flow-through.
  • Add 200ul M-Desulphonation Buffer, wait for 20min, spin at 15,000 rpm for 30sec.
  • Add 200ul M-Wash Buffer, spin for 30 sec.
  • Place the columns into 1.5ml tubes, add 10ul M-Elution Buffer. Wait for 1min, spin at 15,000rpm for 30sec.
  • Measure the DNA with Nanodrop:

RNA