Matt:LabNotes/2014-6-24: Difference between revisions
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>Mzcai (Created page with "==Removal of Amplificiation Adapters== * Lambda digested 13.7ug ssDNA *[[Matt:LabNotes/2014-5-22#Lambda_Exonuclease_...") |
>Mzcai mNo edit summary |
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*Incubated at 94C for 2 min, then 37C for 3 min | *Incubated at 94C for 2 min, then 37C for 3 min | ||
*Added 5ul DpnII | *Added 5ul DpnII | ||
*Incubated at 37C for | *Incubated at 37C for 3 hr | ||
*Added 100ul of the following | *Added 100ul of the following | ||
{| {{table}} | {| {{table}} |
Revision as of 22:58, 24 June 2014
Removal of Amplificiation Adapters
- Lambda digested 13.7ug ssDNA
- Lambda digested 7.87ug ssDNA
- Combine to 300ul of 22ug total ssDNA
To each of 4 x 200ul PCR tubes:
Components | Volume |
ss-amplicon (73.3 ng/ul) | 75 |
USER | 5 |
H2O | 0 |
total | 80 |
*5.5ug 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: 4 tubes of 200ul each