Jie:LabNotes/CpgSeq/2009-5-29

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Bisulfite conversion of PGP DNA for cpg30k ASM validation

No sample sample concentration sample volumn ddH2O conversion reagents conversed DNA concentration and volumn 260:280/260:230
21 PGP1_iPS1 50ng/ul x 1 tubes 20ul 0ul 130ul 179.1ng/ul x 10ul 1.97/1.72
22 PGP1_L 293ng/ul x 1 tubes 6.8ul 13.2ul 130ul 368.7ng/ul x 20ul 2.13/2.04
23 PGP3_L 242ng/ul x 1 tubes 6.8ul 13.2ul 130ul 238.6ng/ul x 20ul 2.07/1.89
24 PGP9_L 249ng/ul x 1 tubes 6.8ul 13.2ul 130ul 223.7ng/ul x 20ul 2.05/1.89
09_8 DF6-9-9_P16(4) 504.8ng/ul x 2 tubes 20ul 0ul 130ul 84ng/ul x 10ul 1.67/1.2
09_9 foreskin 583ng/ul x 2 tube 20ul 0ul 130ul 35.1ng/ul x 10ul 1.2/0.66


  • 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