Jie:LabNotes/CpgSeq/2010-1-22

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Revision as of 18:16, 22 January 2010 by >Jie deng (Created page with '==capture of mouse ES cells from Harvard== ===generation of cpgmm9A=== PCR reaction x 48 x96 H2O …')
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capture of mouse ES cells from Harvard

generation of cpgmm9A

PCR reaction                                            x 48     x96   
H2O                                   49ul             2352ul   4704
EconoTaq 2xMastermix                  50ul             2400ul   4800
Ap1V6IU       (100uM)                0.3ul             14.4ul   28.8   
Ap2V6         (100uM)                0.3ul             14.4ul   28.8
SYBR  green (50x)                    0.4ul             19.2ul   38.4
template(cpgmm9A,10uM)              0.05ul              2.4ul    4.8
Total                                100ul             4800ul   9600
Reaction program: 94c 2min -> (94c 30sec -> 60C 1min -> 72C 45sec)x  13ycles -> 72C 5min -> 15C hold.
I did the ethanol precipitation of PCR amplicon, Lambda exonuclease digestion, UESR and DpnII digestion (with DpnII-oligo_v6). The yield of probe is