Matt:LabNotes/2014-7-22: Difference between revisions

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Line 10: Line 10:
| align="center" style="background:#f0f0f0;"|'''Volume'''
| align="center" style="background:#f0f0f0;"|'''Volume'''
|-
|-
| ss-amplicon (73.3 ng/ul)||75
| ss-amplicon||71.5
|-
|-
| USER||5
| USER||5
|-
|-
| H2O||0
| H2O||3.5
|-
|-
| total||80
| total||80

Revision as of 20:48, 22 July 2014

Agi26k0gap Probe Production Continued

Removal of Amplification Adapters

To each of 7 x 200ul PCR tubes:

Components Volume
ss-amplicon 71.5
USER 5
H2O 3.5
total 80
 *5ug ssDNA per tube
  • Incubated at 37C for 2 hours
  • Added 15ul of the following
Components Volume
10X DpnII Buffer 10
100uM RE-DpnII guide oligo 5
 RE-DpnII guide oligo:
 0-gap: RE DpnII V6
  • Incubated at 94C for 2 min, then 37C for 3 min
  • Added 5ul DpnII
  • Incubated at 37C for 3 hr
  • Added 100ul of the following
Components Volume
10X DpnII Buffer 10
USER 5
H2O 85
Total 100
  • Incubated at 37C for overnight (~15 hrs)

Summary: 7 tubes of 200ul each