Team:Warsaw/Calendar-Main/17 August 2009
From 2009.igem.org
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+ | <h3><div style="text-align: center;">Naking of the plac-RBS-llo part</div></h3> | ||
+ | <h4>Jarek</h4> | ||
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+ | <p>Tasks:</p> | ||
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+ | <li>*Transformating the competent ''E.coli'' strain TopF' with ligation (acquired from Marek) containing RBS-llo gene cloned into vector with plac promoter </li> | ||
+ | <li>*Plating the transformated culure on LB+Amp plates</li> | ||
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Revision as of 07:03, 6 September 2009
Assembly of endosomal detection operon
Marcin
Task 1:
Methods:
- Plasmids were isolated using the A&A plasmid mini kit. Detailed procedure of the isolation is described here
Task 2:
- Digest previously isolated plasmids to verify the correctness of the ligation
Methods:
- Reaction mixture composition:
1 μl purified plasmid DNA product 0.5 μl XbaI (Fermentas) 0.5 μl PstI (Fermentas) 2 μl Buffer Tango (Fermentas) 16.5 μl MQ water
Comment:
All isolated plasmids had the expected construct.
Task 3:
- Prepare bacterial cultures to isolate following constructs:
Task 4:
- Isolate of following constructs:
Methods:
- Plasmids were isolated using the A&A plasmid mini kit. Detailed procedure of the isolation is described here
Task 5:
- Digest previously isolated plasmids to verify the correctness of the ligation
Methods:
- Reaction mixture composition:
1 μl purified plasmid DNA product 0.5 μl XbaI (Fermentas) 0.5 μl PstI (Fermentas) 2 μl Buffer Tango (Fermentas) 16.5 μl MQ water
Comment:
None of isolated plasmids had the expected constructs. Ligations must be prepared once again.
Task 6:
- Restriction digest of following constructs:
Methods:
- Reaction mixture composition:
20 μl purified plasmid DNA product 1 μl XbaI (Fermentas) or 0.5 SpeI (Fermentas) 1 μl PstI (Fermentas) 5 μl Buffer Tango (Fermentas) 24 μl MQ water
Naking of the plac-RBS-llo part
Jarek
Tasks:
- *Transformating the competent ''E.coli'' strain TopF' with ligation (acquired from Marek) containing RBS-llo gene cloned into vector with plac promoter
- *Plating the transformated culure on LB+Amp plates
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