Team:Evry/Protocols/02

From 2013.igem.org

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<h2> Goal </h2>
<h2> Goal </h2>
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<p>In this step, the heat shock (between -80°C up to 42°C) induces bacteria to be in state of competence. DNA from the environment (in our case: plasmid) can then integrate the cell.</p>
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<p>In this step, the heat shock (between -80°C up to 42°C) induces bacteria to be in state of competence. DNA from the environment (in this case: plasmid) can then integrate the cell.</p>
<h2> Preparation </h2>
<h2> Preparation </h2>

Revision as of 09:17, 27 August 2013

Iron coli project

Transformation

Goal

In this step, the heat shock (between -80°C up to 42°C) induces bacteria to be in state of competence. DNA from the environment (in this case: plasmid) can then integrate the cell.

Preparation

For 100 µL of chemical competent cells, add 1 µL of plasmidic DNA. If the ligation protocol is a Golden Gate , add 5 µL instead.
Let 30 minutes on ice.
Let the cells at 42°C for exactly 50 seconds.
Let 5 minutes on ice.
Resuspend the cells with 1 mL of LB spread on a petri dish then let it at 37°C.

Controles

To check if the transformation have work, make a positive and a negative controle.

  • Positive controle: Use plasmid pSB1A3
    If the transformation have worked, colonies must have a red phenotype.

  • Negative controle: Ø
    Without plasmid with antibiotic resistance, no colonies must grow.

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