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Team:Warsaw/Calendar-Main/11 July 2009
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* Alkaline lysis of bacterial cultures to obtain plasmids containing following bricks: | * Alkaline lysis of bacterial cultures to obtain plasmids containing following bricks: | ||
[http://partsregistry.org/Part:BBa_R0010 BBa_R0010] - lacI regulated promoter | [http://partsregistry.org/Part:BBa_R0010 BBa_R0010] - lacI regulated promoter | ||
| - | [http://partsregistry.org/Part:BBa_R0080 | + | [http://partsregistry.org/Part:BBa_R0080 BBa_R0080] - AraC regulated promoter |
* Digestion to confirm plasmid extraction. DNA that should contain pAraC on pSB1A2 plasmid was digested with BamHI PvuI, pLacI on pSB1A2 was digested with IPvuI and PvuII . Digestion mix contained 5ul of extracted DNA, 2ul of Fermentas BamHI buffer , 0.3ul of each enzyme and water added to obtain 20ul total volume. | * Digestion to confirm plasmid extraction. DNA that should contain pAraC on pSB1A2 plasmid was digested with BamHI PvuI, pLacI on pSB1A2 was digested with IPvuI and PvuII . Digestion mix contained 5ul of extracted DNA, 2ul of Fermentas BamHI buffer , 0.3ul of each enzyme and water added to obtain 20ul total volume. | ||
| Line 53: | Line 53: | ||
! expected fragments [bp] | ! expected fragments [bp] | ||
|- | |- | ||
| - | | | + | | placI on pSB1A2 |
| PvuI PvuII | | PvuI PvuII | ||
| 728, 1351 | | 728, 1351 | ||
|- | |- | ||
| - | | | + | | pAraC on pSB1A2 |
| BamHI PvuI | | BamHI PvuI | ||
| 782, 1446 | | 782, 1446 | ||
Revision as of 23:36, 11 July 2009
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Contents |
Gel out phoP/phoQ
Kama
Tasks:
- Isolation of fragment of the correct lenght(˜2200bp)from the gel was performed with the [http://aabiot.com/products/dna_purification/dna_fragments/gel_out/protocol_gel_out.pdf A&A "Gel-out" kit].
Construction of [http://partsregistry.org/wiki/index.php?title=Part:BBa_K177012 K177012] operon1_part2
Ania
Tasks:
- Transformation of chemocompetent strain of E. with [http://partsregistry.org/wiki/index.php?title=Part:BBa_C0012 BBa_C0012 - lacI repressor] on the pSB1A2 ampicillin resistant plasmid taken out of the distribution 200 Kit Plate 1 well 2O.
- Alkaline lysis of bacterial cultures to obtain plasmids containing following bricks:
[http://partsregistry.org/wiki/index.php?title=Part:BBa_R0051 BBa_R0051] - promoter (lambda cI regulated); [http://partsregistry.org/wiki/index.php?title=Part:BBa_B0032 BBa_B0032] - RBS.3 (medium)
- Digestion to confirm plasmid extraction. DNA that should contain R0051 on pSB1A2 plasmid was digested with PvuI and HindII. Digestion mix contained 5ul of extracted DNA, 2ul of Fermentas BamHI buffer , 0.3ul of each enzyme and water added to obtain 20ul total volume.
| DNA sample | restriction enzymes | expected fragments [bp] |
|---|---|---|
| R0051(pcI) on pSB1A2 | PvuI HindII | 695, 1447 |
Creating devices to test promotors in E. coli strains (devices BBa_K177024 and BBa_K177025)
Franek
Tasks:
- Alkaline lysis of bacterial cultures to obtain plasmids containing following bricks:
[http://partsregistry.org/Part:BBa_R0010 BBa_R0010] - lacI regulated promoter [http://partsregistry.org/Part:BBa_R0080 BBa_R0080] - AraC regulated promoter
- Digestion to confirm plasmid extraction. DNA that should contain pAraC on pSB1A2 plasmid was digested with BamHI PvuI, pLacI on pSB1A2 was digested with IPvuI and PvuII . Digestion mix contained 5ul of extracted DNA, 2ul of Fermentas BamHI buffer , 0.3ul of each enzyme and water added to obtain 20ul total volume.
| DNA sample | restriction enzymes | expected fragments [bp] |
|---|---|---|
| placI on pSB1A2 | PvuI PvuII | 728, 1351 |
| pAraC on pSB1A2 | BamHI PvuI | 782, 1446 |
Franek lub Monika opisze minilizy dla Moniki
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