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Team:Warsaw/Calendar-Main/30 September 2009
From 2009.igem.org
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| + | <h3><div style="text-align: center;">Cloning of AraC coding sequence</div></h3> | ||
| + | <h4>Justyna</h4> | ||
| + | <br /> | ||
| + | <p>Task 1:</p> | ||
| + | <ul> | ||
| + | <li><a href="http://partsregistry.org/Part:BBa_B0032">BBa_0032</a> and <a href="http://partsregistry.org/wiki/index.php/Part:BBa_C0080">BBa_C0080</a> digestion</li></ul> | ||
| + | <p>Overnight digestion in 37°C, volume of 30µl<p> | ||
| + | <p><b>Comment:</b></p> | ||
| + | <p>Enzymes: | ||
| + | <br>BBa_B0032 part: SpeI + PstI | ||
| + | <br>BBa_C0080 part: XbaI + PstI</p> | ||
| + | <br><p>Results:</p> | ||
| + | <img src="https://static.igem.org/mediawiki/2009/8/82/B0032C000E0032R000%2BC0040.jpg"></center> | ||
| + | <br><br><br> | ||
| + | <p>Task 2:</p> | ||
| + | <ul> | ||
| + | <li>Ligation of AraC gene and BBa_B0032</li></ul> | ||
| + | <p>4 hour long ligation in room temperature, volume of 25µl<p> | ||
| + | <p>Ligation mix: | ||
| + | <pre> | ||
| + | BBa_B0032 7µl (~49ng) | ||
| + | AraC sequence 15.5µl (~70ng) | ||
| + | buffer 2.5µl | ||
| + | ligase 1µl (Fermentas) | ||
| + | </pre> | ||
| + | <html> | ||
| - | + | <h3><div style="text-align: center;">Fusion of YFP coding sequence and TetR coding sequence</div></h3> | |
| - | + | <h4>Justyna</h4> | |
| + | <br /> | ||
| + | <p>Task 1:</p> | ||
| + | <ul> | ||
| + | <li><a href="http://partsregistry.org/wiki/index.php/Part:BBa_C0040">BBa_C0040</a>+<a href="http://partsregistry.org/Part:BBa_R0080">BBa_R0080 part</a> and <a href="http://partsregistry.org/wiki/index.php/Part:BBa_E0032">BBa_E0032</a> digestion</li></ul> | ||
| + | <p>Overnight digestion in 37°C, volume of 30µl<p> | ||
| + | <p><b>Comment:</b></p> | ||
| + | <p>Enzymes: | ||
| + | <br>BBa_C0040+BBa_R0080 part: SpeI + PstI | ||
| + | <br>BBa_E0032 part: XbaI + PstI</p> | ||
| + | <br><p>Results:</P> above. | ||
| + | <br><br><br> | ||
| + | <p>Task 2:</p> | ||
| + | <ul> | ||
| + | <li>Ligation of YFP gene and BBa_C0040+BBa_R0080</li></ul> | ||
| + | <p>4 hour long ligation in room temperature, volume of 20µl<p> | ||
| + | <p>Ligation mix: | ||
| + | <pre> | ||
| + | BBa_C0040+BBa_R0080 7µl (~50ng) | ||
| + | YFP sequence 9.5µl (~114ng) | ||
| + | buffer 2.0µl | ||
| + | ligase 1µl (Fermentas) | ||
| + | </pre> | ||
| + | </html> | ||
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Revision as of 15:04, 1 October 2009
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Bistable switch testing
Monika
Tasks:
- Transformation of chemocompetent cells (Top10) with plasmid containing bistable switch
Methods:
- Transformation of chemocompetent cells with 10μl of DNA solution - procedure destribed here
- Planting on LB-agar medium supplemented with tetacycline
Results:
Successful, many colonies that express GFP.
Cloning of AraC coding sequence
Justyna
Task 1:
Overnight digestion in 37°C, volume of 30µl
Comment:
Enzymes:
BBa_B0032 part: SpeI + PstI
BBa_C0080 part: XbaI + PstI
Results:
Task 2:
- Ligation of AraC gene and BBa_B0032
4 hour long ligation in room temperature, volume of 25µl
Ligation mix:
BBa_B0032 7µl (~49ng) AraC sequence 15.5µl (~70ng) buffer 2.5µl ligase 1µl (Fermentas)
Fusion of YFP coding sequence and TetR coding sequence
Justyna
Task 1:
- BBa_C0040+BBa_R0080 part and BBa_E0032 digestion
Overnight digestion in 37°C, volume of 30µl
Comment:
Enzymes:
BBa_C0040+BBa_R0080 part: SpeI + PstI
BBa_E0032 part: XbaI + PstI
Results:
above.Task 2:
- Ligation of YFP gene and BBa_C0040+BBa_R0080
4 hour long ligation in room temperature, volume of 20µl
Ligation mix:
BBa_C0040+BBa_R0080 7µl (~50ng) YFP sequence 9.5µl (~114ng) buffer 2.0µl ligase 1µl (Fermentas)
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