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Team:Kyoto/GSDD/Notebook/0907-0913
From 2009.igem.org
- 0727-0802
- Construction.
- 0803-0809
- Construction.
- 0810-0816
- Construction.
- 0817-0823
- Construction.PCR.
- 0824-0830
- Construction.PCR.
- 0831-0906
- Construction.PCR.MPR.
- 0907-0913
- Construction.PCR.MPR.
- 0914-0920
- Construction.PCR.MPR.
- 0921-0927
- Construction.PCR.MPR.
- 0928-1004
- Construction.PCR.MPR.
- 1005-1011
- Construction.PCR.MPR.
- 1012-1018
- Construction.MPR.Observation.
0907-0913 : Construction.PCR.MPR.
Tuesday, 7 September
To Do
- Make repetitive sequence (MPR)
- Make parts(for flush end ligation)
- Annealing
- Restriction Enzyme Digestion
- Ethanol precipitation
(8)
- Ligation
(7)(8)(10)(5)
- transformation
Results
MPR
- Electrophoresis results below indicated that sample E and F are proper for extracting.
Make parts(for flush end ligation)
| NO. | Sample name | conc./(ng/ul) |
|---|---|---|
| 1 | annealing sample | 1.8 |
- This indicated that annealing product was not obtained.
Transformation
- (5)A,(5)B:Colonies were observed.
- (7)A,(7)B,(8),(10)A,(10)B:No colony was observed.
Wednesday, 8 September
To Do
- MPR products (further to 7 Sept.)
- Gel extraction
- Measure concentration
- Make repetitive sequence (MPR)
- (1)(4)(5)
- Colony PCR
- Electrophoresis
- Miniprep and make master plates
Results
MPR products (further to 7 Sept.)
| NO. | band name | conc. |
|---|---|---|
| 1 | 1 | 17.4 |
| 2 | 2 | 19.0 |
| 3 | 3 | 18.4 |
| 4 | 4 | 11.6 |
| 5 | 5 | 17.9 |
Make repetitive sequence (MPR)
- Gel extraction and Measured concentration.
| NO. | sample length /bp | conc. |
|---|---|---|
| 1 | -1000 | 21.5 |
| 2 | 1000-2000 | 42.3 |
| 3 | 2000-5000 | 20.5 |
| 4 | 5000+ | 14.1 |
Colony PCR of (1),(4),(5)
- This result indicates that each part were inserted properly.
Thursday, 9 September
To Do
- PCR products(1)(4)(5)(0908)
- Gel extraction
- Measure concentration (See lab note on 8 Sept.)
- PCR product(5)
- Miniprep and make master plates
- (7)(8)(10)
- Ligation
- Transformation
- Make parts(for flush end ligation) retry
- Annealing
- Restriction Enzyme Digestion
- Ethanol precipitation
Results
(7),(8),(10)
We failed in the transformation 0907 because of the plasmid we used. We tried again using plasmid we purified anew.
- PSB1A2(ES) conc.: 24(ng/ul)
After transformation, colonies were observed in all plates.
Make parts(for flush end ligation) retry
| NO. | sample name | conc. |
|---|---|---|
| 1 | (11) | 49.8 |
| 2 | (11)' | 47.5 |
Friday, 10 August
To Do
- (7)(8)(10)
- Colony PCR
- Electrophoresis
- PCR
Results
(7)(8)(10)
Inserted properly
- (7)A①③,B②
- (8)①
- (10)A①②③,B②③
This result indicates that some samples were inserted properly, though others were self ligated.
PCR
This indicates PCR was achieved.
