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24 August 2008 <|> 26 August 2008
Laboratory work
Team:Input
The following glycerol Stock was made.
[http://partsregistry.org/Part:BBa_J22136 BBa_J22136](insert check OK)
Picked the colony and incubate with 2mL of LB-Ampicillin Medium for 12 hours at 37 degrees.
Took 180μl from culture,and mixed with 880μlLB-Amp Medium.(OD=0.485)
| Liquid culture(OD=0.495) |
0.67μl |
| 60%Glycerol |
0.33μl |
| 20%Glycerol Stock |
1.00ml |
Team:Communication
- (24/8)--->Digestion
- Double Digestion:[http://partsregistry.org/Part:BBa_J04500 BBa_J04500](2007)
- Double Digesiton:[http://partsregistry.org/Part:BBa_R0010 BBa_R0010](2007)
- Single Digestion:[http://partsregistry.org/Part:BBa_R0010 BBa_R0010](2007)
- [http://partsregistry.org/Part:BBa_R0010 BBa_R0010](2007)
| Sample No. |
1 | 2 | 3 | 4 |
| Sample DNA(μL) |
5 | 5 | 1 | 1 |
| EcoRI(μL) |
0.5 | 0.5 | - | - |
| SpeI(μL) |
0.5 | 0.5 | - | - |
| Buffer 1 |
1 | 1 | - | - |
| Buffer 3(μL) |
- | - | 1 | - |
| BSA(μL) |
1 | 1 | - | - |
| dH2O(μL) |
2 | 2 | 7.5 | - |
| TOTAL(μL) |
10 | 10 | 10 | 1 |
- --->Gel Check
|
| Sample No. |
1~3 | 4 |
| Sample DNA(μL) |
10 | 1 |
| Loading Dye(μL) |
2 | 1 |
| dH2O(μL) |
- | 4 |
| TOTAL(μL) |
12 | 6 |
- From right;
- Double Digestion:[http://partsregistry.org/Part:BBa_J04500 BBa_J04500](2007) -> OK
- Double Digesiton:[http://partsregistry.org/Part:BBa_R0010 BBa_R0010](2007) -> ?(low)
- Single Digestion:[http://partsregistry.org/Part:BBa_R0010 BBa_R0010](2007) -> ?(low)
- [http://partsregistry.org/Part:BBa_R0010 BBa_R0010](2007) -> OK(low)
|
--->Digestion
(Double Digestion)
- [http://partsregistry.org/Part:BBa_C0178 BBa_C0178]
- [http://partsregistry.org/Part:BBa_C0170 BBa_C0170]
- [http://partsregistry.org/Part:BBa_J04500 BBa_J04500](2007)
| Sample No. |
1 | 2 | 3 |
| Sample DNA(μL) |
33ng/μl×60μl | 33ng/μl×60μl | 100ng/μl×20μl |
| PstI(μL) |
2 | 2 | 2 |
| XbaI(μL) |
2 | 2 | - |
| SpeI(μL) |
- | - | 2 |
| Buffer 2(μL) |
- | - | 5 |
| Buffer 3(μL) |
10 | 10 | - |
| BSA(μL) |
10 | 10 | 5 |
| dH2O(μL) |
16 | 16 | 16 |
| TOTAL(μL) |
100 | 100 | 52 |
Team:Output
PCR
- [http://partsregistry.org/Part:BBa_E2030 BBa_E2030](1)
- [http://partsregistry.org/Part:BBa_J33202 BBa_J33202](2)
- BL21(3)
- [http://partsregistry.org/Part:BBa_J52008 BBa_J52008](4)
- [http://partsregistry.org/Part:BBa_I712019 BBa_I712019](5)
| No. |
1 | 2 | 3 | 4 | 5 |
| DNA tamplate(μL) |
1 | 1 | 1 | 1 | 1 |
| FW primer(μL) |
5(Venus_YFP_fwd) | 5(LacZ_fwd) | 5(LacZ_fwd) | 5(rLUC_fwd) | 5(fLuc_fwd) |
| VR primer(μL) |
5(VR) | 5(LacZ_rev) | 5(LacZ_rev) | 5(VR) | 5(VR) |
| dNTPmix(μL) |
10 | 10 | 10 | 10 | 10 |
| Thermo pol buffer(μL) |
10 | 10 | 10 | 10 | 10 |
| DNA pol(VENT)(μL) |
1 | 1 | 1 | 1 | 1 |
| dH2O(μL) |
68 | 68 | 68 | 68 | 68 |
| TOTAL(μL) |
100 | 100 | 100 | 100 | 100 |
-->Gel Check
| No. |
1 | 2 | 3 | 4 | 5 |
| DNA tamplate(μL) |
1 | 1 | 1 | 1 | 1 |
| loading Dye(μL) |
1 | 1 | 1 | 1 | 1 |
| dH2O(μL) |
4 | 4 | 4 | 4 | 4 |
| TOTAL(μL) |
6 | 6 | 6 | 6 | 6 |
Mini prep
- [http://partsregistry.org/Part:BBa_E2030 BBa_E2030]
- [http://partsregistry.org/Part:BBa_F2620 BBa_F2620]
Transformation
- [http://partsregistry.org/Part:BBa_E2030 BBa_E2030]
- PUC19
- pGFPUV
- pGEX Venus YFP
- Membren Venus YFP
PCR
- [http://partsregistry.org/Part:BBa_J63001 BBa_J63001](1)
| No. |
1 |
| DNA tamplate(μL) |
1 |
| FW primer(Venus)(μL) |
5 |
| VR primer(μL) |
5 |
| dNTPmix(μL) |
10 |
| Thermo pol buffer(μL) |
10 |
| DNA pol(VENT)(μL) |
1 |
| dH2O(μL) |
68 |
| TOTAL(μL) |
100 |
"
