Difference between revisions of "Team:Paris Bettencourt/Notebook/Phytase"
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<br><h1 class="date one">August 30th</h1> | <br><h1 class="date one">August 30th</h1> | ||
<h2>Transformation on yeasts with <i>Cre</i> gene</h2> | <h2>Transformation on yeasts with <i>Cre</i> gene</h2> | ||
+ | First step, we realize the PCR of CRE and RFP gene with primers (created by Amaury) and plasmid containing RFP. We amplified the gene, its promoter and its terminater.<br> | ||
+ | <b>Protocol</b> :<a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols/PCR"> PCR</a><br> | ||
+ | <br> | ||
+ | |||
+ | Second step, we make PCR purification, and check the result with electrophoresis.<br> | ||
+ | <b>Protocol</b> :<a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols/PCR_purification"> PCR purification</a> | ||
+ | <br> | ||
+ | |||
+ | Third step, we transformed yeast that is already integrated the resistance instead of PHO85 gene. We introduce the CRE and RFP gene instead of PHO80 gene<br> | ||
+ | <b>Protocol</b> :<a href="https://2015.igem.org/Team:Paris_Bettencourt/Protocols/Heat_shock_transformation_for_yeast"> Transformation of yeast</a><br> | ||
+ | |||
+ | Fourth step, we cultived the transforming yeasts in YPD agar medium with geneticin overnight. | ||
− | We watching any culture on plates YPD + Geneticin. The yeast are not resistant to geneticin, | + | We watching any culture on plates YPD + Geneticin. The yeast are not resistant to geneticin, they had not introduced the resistance gene. Transformation does not work.</p> |
<h2>PCR and result</h2> | <h2>PCR and result</h2> | ||
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We restart the transformation of September 2nd, with Cre sequence and RFP gene.<br> | We restart the transformation of September 2nd, with Cre sequence and RFP gene.<br> | ||
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<h2> Analysis of results of titration phytic acid of September 2nd</h2> | <h2> Analysis of results of titration phytic acid of September 2nd</h2> |
Revision as of 13:11, 18 September 2015