AlanFung:LabNotes/Sequencing/2009-11-24: Difference between revisions

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>Alan6017518
>Alan6017518
Line 13: Line 13:
| Amp R6.3NH2 (100uM)||0.2
| Amp R6.3NH2 (100uM)||0.2
|-
|-
| dUTP (1mM)||0.4
| 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

End Repair