Team:Queens Canada/Notebook/Protocols/HeatShock

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<classred1> <b> Rehydration </b></classred1>
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    <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/Rehydration">rehydration      </a></span>     </regulartext>
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    <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/HeatShock">Heat Shock      </a></span>     </regulartext>
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<classred1> <b> heat shock </b></classred1>
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     <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/LiquidCulture">Liquid Culture     </a></span>    </regulartext>
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     <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/LiquidCulture">liquid culture     </a></span>    </regulartext>
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     <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/GlycerolStock">glycerol stock     </a></span>    </regulartext>
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     <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/Miniprep">Miniprep     </a></span>    </regulartext>
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     <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/Miniprep">miniprep     </a></span>    </regulartext>
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     <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/Digestion">digestion     </a></span>    </regulartext>
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     <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/Ligation">Ligation     </a></span>    </regulartext>
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     <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/Ligation">ligation     </a></span>    </regulartext>
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     <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/GelElectrophoresis">Electrophoresis     </a></span>    </regulartext>
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     <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/Extraction"> Extraction     </a></span>    </regulartext>
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     <span class="classred"><a href="https://2011.igem.org/Team:Queens_Canada/Notebook/Protocols/Extraction"> extraction     </a></span>    </regulartext>
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<h3red> Rehydration- iGEM Standard Parts</h3red>
 
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<h3red> Rehydration- Primers</h3red>
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<h3red> Heat Shock (Transformation) </h3red> <p>
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<regulartext> <b> Storage and Labelling </b> </regulartext> <br>
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<regulartext> - Store the transformed cells in the 4°C fridge. </regulartext> <br>
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<regulartext> - Label the product plate as “EC (resistance)” and in accordance with the standard labelling format, as outlined at the front of your lab book and on Google Docs. </regulartext> <p>
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<regulartext> <b> Materials </b> </regulartext> <br>
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<regulartext> - 100μL of chemical competent cells per transformation  </regulartext> <br>
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<regulartext> - Miniprep plasmid DNA </regulartext> <br>
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<regulartext> -2XTY or SOC medium </regulartext> <br>
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<regulartext> - Antibiotic plates (according to plasmid) </regulartext> <p>
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<regulartext> <b> Procedure </b> </regulartext> <br>
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<regulartext> 1. Take out competent cells from -80⁰C and put them on ice immediately before they are needed. <br>
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2. Add 2μl plasmid DNA to thawed cells and mix by flicking the side of the tube. <br>
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3. Incubate on ice for 20 minutes. <br>
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4. Pre-warm antibiotic plates in 37⁰C incubator. <br>
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5. Heat shock for 1 min 15 sec at 42⁰C.<br>
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6. Place on Ice for 2 minutes. <br>
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7. Add 500ul 2XTY (or SOC) medium (kept at room temp) to each tube (medium with NO antibiotic). <br>
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8. Shake the tubes at 37⁰C for 1 hour on a shaking incubator. <br>
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9. Spread 100μl of each transformation tube on appropriate antibiotic plates.  <br>
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-In addition, you can: Take 1μL of TOP10 and add 99μL of growth buffer, plate it. Spin remainder for 1 min at 13000, discard supernatant, re-suspend in 100μL of growth buffer<br>
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10. Incubate at 37⁰C overnight.<br>
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</regulartext> <br>
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Latest revision as of 20:32, 28 September 2011

Queen's
heat shock
Heat Shock (Transformation)

Storage and Labelling
- Store the transformed cells in the 4°C fridge.
- Label the product plate as “EC (resistance)” and in accordance with the standard labelling format, as outlined at the front of your lab book and on Google Docs.

Materials
- 100μL of chemical competent cells per transformation
- Miniprep plasmid DNA
-2XTY or SOC medium
- Antibiotic plates (according to plasmid)

Procedure
1. Take out competent cells from -80⁰C and put them on ice immediately before they are needed.
2. Add 2μl plasmid DNA to thawed cells and mix by flicking the side of the tube.
3. Incubate on ice for 20 minutes.
4. Pre-warm antibiotic plates in 37⁰C incubator.
5. Heat shock for 1 min 15 sec at 42⁰C.
6. Place on Ice for 2 minutes.
7. Add 500ul 2XTY (or SOC) medium (kept at room temp) to each tube (medium with NO antibiotic).
8. Shake the tubes at 37⁰C for 1 hour on a shaking incubator.
9. Spread 100μl of each transformation tube on appropriate antibiotic plates.
-In addition, you can: Take 1μL of TOP10 and add 99μL of growth buffer, plate it. Spin remainder for 1 min at 13000, discard supernatant, re-suspend in 100μL of growth buffer
10. Incubate at 37⁰C overnight.

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