Kun:LabNotes/ASE/2007-12-6
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Exp.1 PCR on the pooled captured DNA
I still have some capturing reactions from 11-21-07, 11-26-07, 12-03-07 left. I decided to pool the reactions from the same templates togethers, and do PCR to make more Solexa sequencing library. The reason is that, based on the quantification experiment did yesterday, there are still not enough number of circularized molecules per target.
one reaction 8 reactions Template: 5~10ul each 10x Buffer: 9.0ul 72ul 25mM MgCl2 5.4ul 43.2ul 25mM dNTP 1.2ul 9.6ul 100uM AmpSolV6F2 0.4ul 3.2ul 100uM AmpSolV6R2 0.4ul 3.2ul iTaq 1ul 8ul 50X SYBG I 0.8ul 6.4ul H2O 72ul 576ul
94C 3min -> 10 cycles of (94C 45sec -> 57C 1min -> 72C 1min) -> 12 cycles of (94C 45 sec -> 72 1min) -> 72C 3min -> 15C hold.