Jie:LabNotes/CpgSeq/2008-8-29: Difference between revisions

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>Jie deng
(New page: ==3rd Bisulfite conversion of DNA using Zymo EZ DNA Methylation-Gold kit== {| border="1" cellpadding="5" cellspacing="0" align="center" |- | align="center" style="background:#f0f0f0;"|'''...)
 
>Jie deng
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==3rd Bisulfite conversion of DNA using Zymo EZ DNA Methylation-Gold kit==


{| border="1" cellpadding="5" cellspacing="0" align="center"
|-
| align="center" style="background:#f0f0f0;"|''' '''
| align="center" style="background:#f0f0f0;"|'''sample '''
|align="center" style="background:#f0f0f0;"|'''sample concentration'''
| align="center" style="background:#f0f0f0;"|'''sample volumn'''
| align="center" style="background:#f0f0f0;"|'''ddH2O '''
| align="center" style="background:#f0f0f0;"|'''conversion reagents'''
| align="center" style="background:#f0f0f0;"|'''conversed DNA concentration and volumn'''
| align="center" style="background:#f0f0f0;"|'''260:280/260:230'''
|-
|1||BJ||178ng/ul||11.2ul||8.8ul||130ul||ng/ul x 10ul||2.18/2.97
|-
|2||BJ_iPS_11||140ng/ul||14.3ul||5.7ul||130ul||ng/ul x 10ul||2.19/2.91
|-
|3||BJ_iPS_12||40ng/ul||20ul||0ul||130ul||ng/ul x 10ul
|-
|9||Hybrid2||200ng/ul||10ul||10ul||130ul||ng/ul x 10ul||2.3/3.0
|-
| ||Jurkat||200ng/ul x 2 tube||10ul||10ul||130ul||ng/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

Latest revision as of 16:24, 29 August 2008