AlanFung:LabNotes/Capturing/2014-1-14: Difference between revisions
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==Bisulfite Conversion of Jurkat gDNA== | ==Bisulfite Conversion of Jurkat gDNA== | ||
*Bisulfite conversion with zymo methylation gold | *Bisulfite conversion with zymo methylation gold | ||
*Convert 500ng of Jurkat gDNA | *Convert 500ng of Jurkat gDNA (2 rxn, 100ng total) | ||
*Add 130ul of lighting conversion reagent to 5ul of jurkat gDNA and 15ul of ddH2O | *Add 130ul of lighting conversion reagent to 5ul of jurkat gDNA and 15ul of ddH2O | ||
*Mix and centrifuge | *Mix and centrifuge |
Revision as of 01:30, 15 January 2014
Capturing Bisulfite Converted Jurkat gDNA w GP1 V6 probe set
Capturing Jurkat gDNA w ppMALT1
Bisulfite Conversion of Jurkat gDNA
- Bisulfite conversion with zymo methylation gold
- Convert 500ng of Jurkat gDNA (2 rxn, 100ng total)
- Add 130ul of lighting conversion reagent to 5ul of jurkat gDNA and 15ul of ddH2O
- Mix and centrifuge
1. 98°C for 8 minutes 2. 54°C for 60 minutes 3. 4°C hold
- Add 600ul M-binding buffer to column
- load the sample into column, mix by inverting column and centrifuge at 14000rpm for 30sec
- discard flow through
- add 100ul m-wash buffer to the column and centrifuge at 14000rpm for 30sec
- add 200ul L-Desulphonation buffer to the column and sit at RT for 20 minutes and centrifuge at 14000rpm for 30sec
- add 200ul of m-wash buffer to the column and centrifuge at 14000rpm for 30sec
- Add 10ul of EBuffer to column sit for 5 mintues and centrifuge at 14000rpm for 30sec (2 rxn total)
ssDNA Qubit assay
47.1ng/ul *20ul = 942ng