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.