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Revision as of 08:39, 16 July 2008
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Preparation of Competent Cells
- Dilution 1/200e (1mL of the O/N culture on 200mL of LB)
- Culture at 37°C until DO=0,57 (around 3h)
- Sharing into 4 Falcon tubes of 50mL and cooling at 4°C during 40' in ice
- Spin during 8', 2000rpm, 4°C
- Resuspension of the cell's pellet in 20mL of cold CaCl2 (0,1M) during 30' in ice
- Spin during 8', 2000rpm, 4°C
- Resuspension of the cell's pellet in 1mL of cold CaCl2 (0,1M)
- 1h of delay unexpected (problem with DNA lyophilized)
- Transformation of the Biobricks with the "classical" protocol (--> heat shock at 42°C during 40")
- Incubation O/N at 37°C of the transformed bacteria
Remark :
- Biobricks 2007 : use of 3µl of the minipreps
- Biobricks 2008 : use of the maximum recoverable volume of the minipreps
Liste of the transformation:
| Miniprep
| Details
| Results
|
| MP1
| J23100
| Transformation
|
| MP2
| J23107
| Transformation
|
| MP3
| B0015
| Transformation
|
| MP5
| JB0030
| Transformation
|
| MP6
| E0422
| Transformation
|
| Ref. Book
| Plasmid
| Antibio
| Plate 2007
| Results
|
| 2E_SP 1000
| B0034
| A
| 3O P1
| No transformation
|
| 10F_SP 1001
| R0079
| A
| 19N P1
| No transformation
|
| 4F_SP 1013
| C0079
| K
| 7K P2
| No transformation
|
| 6H_SP 1001
| C0179
| A
| 6H P1
| No transformation
|
| 6D_SP 1007
| S03879
| A
| /
| No transformation
|
| 6B_SP 1001
| S03154
| A
| 24K P1
| No transformation
|
| 10F_SP 1002
| J23101
| A
| 21G P3
| No transformation
|
| 11F_SP 1002
| J23109
| A
| 23G P3
| No transformation
|
| 4C_SP 1000
| R0040
| A
| 7O P1
| No transformation
|
| 2G_SP 1000
| C0040
| A
| 5C P1
| No transformation
|
| 1C_SP 1008
| PSB1A3
| A
| 23E P1
| No transformation
|
| 7G_SP 1013
| E1010
| K
| 15M P2
| No transformation
|
| 7E_SP 1001
| E0040
| A
| 5H P1
| No transformation
|
| 2F_SP 1016
| E0030
| K
| 7G P1
| No transformation
|
Stock solution :
- TE (10:1) pH 8.0
- plates of Ampicilin and Kanamycin
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