Matt:LabNotes/2014-7-22: Difference between revisions
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>Mzcai (Created page with "==Agi26k0gap Probe Production Continued== ===Removal of Amplification Adapters=== *[[Matt:LabNotes/2014-7-1#Lambda_Exonuclease_Digestion | Lambda digested 14.2ug (200ul) ssDNA...") |
>Mzcai mNo edit summary |
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*[[Matt:LabNotes/2014-7-1#Lambda_Exonuclease_Digestion | Lambda digested 14.2ug (200ul) ssDNA]] | *[[Matt:LabNotes/2014-7-1#Lambda_Exonuclease_Digestion | Lambda digested 14.2ug (200ul) ssDNA]] | ||
*[[Matt:LabNotes/2014-7-17#Lambda_Exonuclease_Digestion | Lambda digested 21.4ug (320ul)ssDNA]] | *[[Matt:LabNotes/2014-7-17#Lambda_Exonuclease_Digestion | Lambda digested 21.4ug (320ul)ssDNA]] | ||
*Combine to | *Combine to 520ul of 35ug total ssDNA | ||
To each of 7 x 200ul PCR tubes: | To each of 7 x 200ul PCR tubes: |
Revision as of 18:24, 22 July 2014
Agi26k0gap Probe Production Continued
Removal of Amplification Adapters
- Lambda digested 14.2ug (200ul) ssDNA
- Lambda digested 21.4ug (320ul)ssDNA
- Combine to 520ul of 35ug total ssDNA
To each of 7 x 200ul PCR tubes:
Components | Volume |
ss-amplicon (73.3 ng/ul) | 75 |
USER | 5 |
H2O | 0 |
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