Biomod/2011/LMU/FolD'N'Assemble/Protocols: Difference between revisions
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===06-03-11(Timon)=== | ===06-03-11(Timon)=== | ||
[[Image:20110603 1.i-motif.JPG|thumb|upright=1.5|06-03-11,left to right, Ladder 1kb, Scaffold, momomers, dimers(4b), dimers with i-motif]] | [[Image:20110603 1.i-motif.JPG|thumb|upright=1.5|06-03-11,left to right, Ladder 1kb, Scaffold, momomers, dimers(4b), dimers with i-motif]] | ||
Ran a gel with 3 samples | Ran a gel(2% agaroses) with 3 samples | ||
*monomers(cs) | *monomers(cs) | ||
*dimers(cs+4es) | *dimers(cs+4es) | ||
*dimers + i-motif (cs+i-m) | *dimers + i-motif (cs+i-m) |
Revision as of 05:39, 5 June 2011
May
05-13-11(Timon, Alex, Ralf)
We wanted to find the best MgCl concentration and the best annealing time for our construct.
SampleNr. | Annealing time | MgCl(mM) | |
---|---|---|---|
1 | 4h | 10 | |
2 | 4h | 14 | |
3 | 4h | 18 | |
4 | 24h | 10 | |
5 | 24h | 14 | |
6 | 24h | 18 | |
7 | 24h | 14 | dimers(2b) |
8 | 55h | 10 | |
9 | 55h | 14 | |
10 | 55h | 18 |
Then we used electrophoresis in 2% agarose gel to test the samples
05-16-11 (Timon, Alex, Ralf)
Another test for best MgCl concentration and annealing time. We used a 2% agarose gel.
SampleNr. | Annealing time | MgCl(mM) | |
---|---|---|---|
1 | 55h | 16 | |
2 | 55h | 18 | |
3 | 55h | 16 | dimers(2b) |
4 | 55h | 18 | dimers(2b) |
5 | 55h | 16 | dimers(4b) |
6 | 55h | 18 | dimers(4b) |
05-17-11(Timon, Alex, Ralf)
We took first TEM (transmission electron microscope) pictures of our construct.
05-18-11(Timon, Alex, Ralf)
New samples for next week: 200μL, 18mM MgCl, 55h
05-23-11(Timon, Alex, Ralf)
We prepared buffers with different pH.
- pH 4,5
- pH 5
- pH 6
- pH 8
05-24-11(Alex, Ralf, Timon)
- prepared 3 samples of origamis for measuring its stability in a changing pH environment
- exchanged standard buffer with pH adjusted buffer using centrifugal filters
- 3 samples, ~24 h exposure to pH values of 4.5, 5 and 6, 1x TE buffer, 18mM MgCl2
05-25-11(Alex)
- made a gel with pH adjusted buffers, UV screenshot inconclusive, going to repeat experiment on Mo 05-30-11
05-27-11(Alex)
- prepared samples to repeat the buffer pH adjusted gel, 150 μL, standard recipe
05-30-11(Alex)
- ran a gel with 5 samples (2% Agarose)
- pH 8 (control sample, no buffer exchange at all)
- pH 8 (no pH change but performed the buffer exchange procedure)
- pH 6
- pH 5
- pH 4.5
- origami seems stable under changing pH conditions, extracted sample with pH 5 for further TEM analysis
05-31-11(Timon)
- made new staple mix for cs 200nM
- started annealing new samples with i-motif
06-03-11(Timon)
Ran a gel(2% agaroses) with 3 samples
- monomers(cs)
- dimers(cs+4es)
- dimers + i-motif (cs+i-m)