Kun:LabNotes/CpgSeq/2007-12-21
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Perform a capturing experiment using bisulfite-PCR product as a control[edit]
Pooled PCR product from bisulfite converted Jurkat genomic DNA: 19ng/ul -> 100nM.
JK bis-DNA: 0ul 0ul 10ul 10ul JK-PCR amplicon: (10nM) 1ul (0.1nM) 1ul (0.1nM) 1ul 0ul 10X buffer 1.5ul 1.5ul 1.5ul 1.5ul 10nM CpG-MIP v2 2ul 2ul 2ul 2ul H2O 10.5ul 10.5ul 0.5ul 1.5ul
95C 3min->60C 42h-> add 1ul SLN mix (1U/ul stoffel, 1U/ul AmpLigase, 200uM dNTP) -> 65C 1h -> 20 cycles of (95C 1min -> 65C 1h) -> 65C 2h -> 37C 1min -> add 1ul of exo mix (Exo I, Exo III, T7 exo), 0.5ul USER enzyme->37c 1 hour -> 94C 5min -> 4C hold.