Daniel:Notebook/HiResChrPaint/2013-8-2: Difference between revisions
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5. [[Daniel:Protocols/Qiaquick_Column|Column Purification]], elute with 40 uL EB | 5. [[Daniel:Protocols/Qiaquick_Column|Column Purification]], elute with 40 uL EB | ||
6. Measure concentrations in nanodrop | 6. Measure concentrations in nanodrop | ||
===qPCR Results=== | |||
===Nanodrop Results=== | |||
==DpnII Digestion== |
Revision as of 19:44, 2 August 2013
S1 Probe Preparation
Armed with a new bottle of water (seriously...), I'm now ready to amplify the S1 set.
PCR Amplification
1. Reaction Table
20 nM OligoPool | 2X Kapa SYBR qPCR MM | 100 uM AP1V41U | 100 uM AP2V4 | 100 uM AP1V61U | 100 uM AP2V6 | 2 mM aa-dUTP | H20 | Total Volume | |
per rxn | 0.2 | 50 | 0.4 | 0.4 | 0.4 | 0.4 | 2.5 | 45.7 | 100 |
V4S1 (12.5x) | 2.5 | 625 | 5 | 5 | 0 | 0 | 31 | 581.5 | 1250 |
V6S1 (12.5x) | 2.5 | 625 | 0 | 0 | 5 | 5 | 31 | 581.5 | 1250 |
2. Aliquot 100 uL into each of 12 qPCR tubes 3. PCR protocol: i. PCR 3 min 95C ii. 95C 3 sec iii. 55C 30 sec iv. 72C 20 sec v. plate read vi. goto ii x17 vii. 72C 2 min viii. 16C hold 4. Pool 3 samples together each (300 uL each, 4 pools per set, 8 total) 5. Column Purification, elute with 40 uL EB 6. Measure concentrations in nanodrop