Difference between revisions of "Team:BOKU-Vienna/Protocol"
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| + | <li class="small"><a href="#transformation">Transformation of chemically competent E. Coli</a></li> | ||
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<h3 id="methods">Methods</h3> | <h3 id="methods">Methods</h3> | ||
| − | <h4 id=" | + | <h4 id="transformation">Transformation of chemically competent E. Coli</h4> |
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| + | <p> Competent cells from E. Coli strain DH5α or DH10B were thawed on ice. The whole ligation/other DNA samples were added to the cells, for 30 min the mix was put on ice. The cells and DNA mix were heat shocked for 90s at 42°C. The cells recovered on ice for 5min. 1 mL of LB medium was added to the cells and then followed an incubation at 37°C. | ||
| + | shaker-incubator (150 rpm) for 45 min to 1 h. Plate the appropriate amount of cells on selective LB agar.</p> | ||
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| + | <h4 id="miniprep">Miniprep</h4> | ||
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<p>Minipreps were performed using Hi Yield®Plasmid Mini DNA Isolationkit according to the manufacturer's protocols. | <p>Minipreps were performed using Hi Yield®Plasmid Mini DNA Isolationkit according to the manufacturer's protocols. | ||
</p> | </p> | ||
Revision as of 13:39, 10 July 2017
Protocol
Protocols from A-Z.
Here you will find our protocols in the future.
Contents
Materials
LB medium - E. Coli
| Name | Amount |
|---|---|
| Hy Soy Peptone | 10.0g |
| Yeast Extract | 5.0g |
| NaCl | 5.0g |
| RO-water | 1000 mL |
LB Agar - E. Coli
| Name | Amount |
|---|---|
| Hy soy Peptone | 10.0g |
| Yeast Extract | 5.0g |
| NaCl | 5.0g |
| RO-water | 980 mL |
| Agar-Agar per 500mL | 10g |
Methods
Transformation of chemically competent E. Coli
Competent cells from E. Coli strain DH5α or DH10B were thawed on ice. The whole ligation/other DNA samples were added to the cells, for 30 min the mix was put on ice. The cells and DNA mix were heat shocked for 90s at 42°C. The cells recovered on ice for 5min. 1 mL of LB medium was added to the cells and then followed an incubation at 37°C. shaker-incubator (150 rpm) for 45 min to 1 h. Plate the appropriate amount of cells on selective LB agar.
Miniprep
Minipreps were performed using Hi Yield®Plasmid Mini DNA Isolationkit according to the manufacturer's protocols.
