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Team:Kyoto/GSDD/Notebook/0831-0904
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.
0831-0904 : what to do. what to do. what to do. what to do.
Monday, 31 August
To Do
- Miniprep p2-1-1, p2-1-2, p2-4-2
- Transformation p2-4-1(retry)
- MPR
Results
Miniprep
- p2-1-1, p2-1-2, p2-4-2 was minipreped.
| NO. | Sample name | conc./ul |
|---|---|---|
| 1 | p2-1-1 | 67.9 |
| 2 | p2-1-2 | 61.5 |
| 3 | p2-4-2 | 94.4 |
Transformation
- p2-4-1 was transformed, and colony was observed.
MPR
- xxx was MPRed.
| NO. | 1 | 2 | 3 | 4 | 5 | 6 |
|---|---|---|---|---|---|---|
| temp.X deg C | 60 | 60 | 60 | 60 | 60 | 60 |
| MPR primer 1 | Fw1 | Fw2 | Fw3 | Fw1 | Fw2 | Fw3 |
| MPR primer 2 | Rev1 | Rev1 | Rev1 | Rev2 | Rev2 | Rev2 |
| NO. | 1 | 2 | 3 | 4 | 5 | 6 |
| temp.X deg C | 63 | 63 | 63 | 63 | 63 | 63 |
| MPR primer 1 | Fw1 | Fw2 | Fw3 | Fw1 | Fw2 | Fw3 |
| MPR primer 2 | Rev1 | Rev1 | Rev1 | Rev2 | Rev2 | Rev2 |
| NO. | 1 | 2 | 3 | 4 | 5 | 6 |
| temp.X deg C | 66 | 66 | 66 | 66 | 66 | 66 |
| MPR primer 1 | Fw1 | Fw2 | Fw3 | Fw1 | Fw2 | Fw3 |
| MPR primer 2 | Rev1 | Rev1 | Rev1 | Rev2 | Rev2 | Rev2 |
- Bands were observed on samples with Fw2.
- Bands were longer on samples with Rev1 than with Rev2.
Tuesday, 1 September
To Do
- Electrophoresis of MPR produnts on 31 Aug.
- Miniprep
Results
Electrophoresis
(See lab-note on 31 Aug.)
Wednesday, 2 September
To Do
- Miniprep of p2-4-1
- PCR of (7), (8), (10)
- PCR of (7), (10) (retry)
Results
Miniprep
- p2-4-1 was minipreped.
| NO. | Sample name | conc./ul |
|---|---|---|
| 1 | p2-4-1 | 92.1 |
PCR
- (7), (8), (10) was PCRed.
| NO. | Sample name | conc./(ng/ul) |
|---|---|---|
| 1 | (7) | 9.7 |
| 2 | (8) | 73.2 |
| 3 | (10) | 6.9 |
- (7), (10) was PCRed twice (again).
| NO. | Sample name | conc./(ng/ul) |
|---|---|---|
| 1 | (7)A | 2.9 |
| 2 | (7)B | 9.3 |
| 3 | (10)A | 5.5 |
| 4 | (10)B | 9.1 |
These concentration suggest that PCR was not achieved. Low concentration of primer causes PCR fault. We should have mix well primer when we add water in primer tube. We are going to retry PCR of (7),(10) the next day.
Thursday, 3 September
To Do
- measuring concentration of (7), (10) (see lab-note on 2 Sept.)
- PCR and measuring concentration of (7), (10) (retry)
- Restriction Enzyme, and PCR purification (8).
Results
Electrophoresis of PCR (7), (10)
Concentration of PCR (7), (10)
| NO. | Sample name | conc./(ng/ul) |
|---|---|---|
| 1 | (7)A | 71.7 |
| 2 | (7)B | 104.3 |
| 3 | (10)A | 33.5 |
| 4 | (10)B | 42.6 |
Friday, 4 September
To Do
(5)
- PCR
- PCR purification kit
(5), (7), (10)
- Restriction Enzyme
- purification kit
- ligation
Results
Electrophoresis of PCR (5)
Concentration of PCR (5)
| NO. | Sample name | conc./(ng/ul) |
|---|---|---|
| 1 | (5)A | 143.4 |
| 2 | (5)B | 174.8 |
Concentration of PCR prification kit (5), (7), (8), (10)
| NO. | Sample name | conc./(ng/ul) |
|---|---|---|
| 1 | (5)A | 53.9 |
| 2 | (5)B | 95.4 |
| 3 | (7)A | 28.9 |
| 4 | (7)B | 33.3 |
| 5 | (8) | 68.3 |
| 6 | (10)A | 29.0 |
| 7 | (10)B | 32.2 |
