AlanFung:LabNotes/Sequencing/2009-11-24: Difference between revisions
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| Amp R6.3NH2 (100uM)||0.2 | | Amp R6.3NH2 (100uM)||0.2 | ||
|- | |- | ||
| dUTP (1mM)|| | | dUTP (1mM)||2 | ||
|- | |- | ||
| 50X SYBG I||0.4 | | 50X SYBG I||0.4 |
Revision as of 19:36, 24 November 2009
Sequencing Library Construction with Single Adaptor Protocol-CVF-N Set
- To make 100ng of CVF-N set (A-D)
PCR with dUTP and 6.3NH2 Primer
1 well | |
H2O | 46.6 |
Amp F6.3NH2(100uM) | 0.2 |
Amp R6.3NH2 (100uM) | 0.2 |
dUTP (1mM) | 2 |
50X SYBG I | 0.4 |
2X Master Mix | 50 |
Template | 1 |
Total | 100 |
- Qiaquick purification, elute with 32ul
USER Digestion
DNA | 30 |
USER | 3 |
Total | 33 |
S1 Nuclease Digestion
S1 Digestion | |
10X S1 Nuclease Buffer | 4 |
DNA after USER Digestion | 33 |
S1 Nuclease (10U/ul) | 1 |
ddH2O | 2 |
- 37C 10mins
- Minelute purification. Elute w/16uH2O