Jie:LabNotes/CpgSeq/2009-5-29: Difference between revisions

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| align="center" style="background:#f0f0f0;"|'''conversion reagents'''
| align="center" style="background:#f0f0f0;"|'''conversion reagents'''
| align="center" style="background:#f0f0f0;"|'''conversed DNA concentration and volumn'''
| align="center" style="background:#f0f0f0;"|'''conversed DNA concentration and volumn'''
| align="center" style="background:#f0f0f0;"|'''260:280/260:230'''
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|20||GM20431||256ng/ul x 1 tube||8ul||12ul||130ul||
|20||GM20431||256ng/ul x 1 tube||8ul||12ul||130ul||56.1ng/ul x 10ul
|-
|-
|21||PGP1_iPS1||50ng/ul x 1 tubes||10ul||10ul||130ul||179.1ng/ul x 10ul||1.97/1.72
|21||PGP1_iPS1||50ng/ul x 1 tubes||10ul||10ul||130ul||40.7ng/ul x 10ul
|-
|-
|22||PGP1_L||293ng/ul x 1 tubes||6.8ul||13.2ul||130ul||368.7ng/ul x 20ul||2.13/2.04
|22||PGP1_L||293ng/ul x 1 tubes||6.8ul||13.2ul||130ul||191.6ng/ul x 10ul
|-
|-
|23||PGP3_L||242ng/ul x 1 tubes||6.8ul||13.2ul||130ul||238.6ng/ul x 20ul||2.07/1.89
|23||PGP3_L||242ng/ul x 1 tubes||6.8ul||13.2ul||130ul||114.7ng/ul x 10ul
|-
|-
|24||PGP9_L||249ng/ul x 1 tubes||6.8ul||13.2ul||130ul||223.7ng/ul x 20ul||2.05/1.89
|24||PGP9_L||249ng/ul x 1 tubes||6.8ul||13.2ul||130ul||106.5ng/ul x 10ul
|-
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|09_8||DF6-9-9_P16(4)||504.8ng/ul x 3 tubes||4ul||16ul||130ul||84ng/ul x 10ul||1.67/1.2
|09_8||DF6-9-9_P16(4)||504.8ng/ul x 3 tubes||4ul||16ul||130ul||285.7ng/ul x 10ul||1.67/1.2
|-
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|09_9||foreskin||583ng/ul x 2 tube||4ul||16ul||130ul||35.1ng/ul x 10ul||1.2/0.66
|09_9||foreskin||583ng/ul x 2 tube||4ul||16ul||130ul||212.5ng/ul x 10ul
|-
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|}
|}

Revision as of 21:44, 29 May 2009

Bisulfite conversion of PGP DNA for cpg30k ASM validation

No sample sample concentration sample volumn ddH2O conversion reagents conversed DNA concentration and volumn
20 GM20431 256ng/ul x 1 tube 8ul 12ul 130ul 56.1ng/ul x 10ul
21 PGP1_iPS1 50ng/ul x 1 tubes 10ul 10ul 130ul 40.7ng/ul x 10ul
22 PGP1_L 293ng/ul x 1 tubes 6.8ul 13.2ul 130ul 191.6ng/ul x 10ul
23 PGP3_L 242ng/ul x 1 tubes 6.8ul 13.2ul 130ul 114.7ng/ul x 10ul
24 PGP9_L 249ng/ul x 1 tubes 6.8ul 13.2ul 130ul 106.5ng/ul x 10ul
09_8 DF6-9-9_P16(4) 504.8ng/ul x 3 tubes 4ul 16ul 130ul 285.7ng/ul x 10ul 1.67/1.2
09_9 foreskin 583ng/ul x 2 tube 4ul 16ul 130ul 212.5ng/ul x 10ul


  • 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