AlanFung:Protocol/Construction of Solexa sequencing Library(USER): Difference between revisions
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==S1 Nuclease Digestion== | ==S1 Nuclease Digestion== | ||
{| {{table}} | |||
| 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 | |||
==End Repair== |
Revision as of 16:50, 24 November 2009
Sequencing Library Construction with Single Adaptor Protocol
PCR with dUTP and 6.3NH2 Primer
1 well | |
H2O | 47.4 |
Amp F6.3NH2(100uM) | 0.2 |
Amp R6.3NH2 (100uM) | 0.2 |
dUTP (1mM) | 0.8 |
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