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| - | {{:Team:TUDelft/Scripts}}{{:Team:TUDelft/CSS}}{{:Team:TUDelft/menu}}
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| - | <html>
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| - | <head><title>Receptor</title></head>
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| - | <body>
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| - | <div style="height:70px; width:100%;"></div>
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| - | <div id="logo_ed"><a href="https://2012.igem.org/Team:TU-Delft" 'onfocus=this.blur()'><img src="https://static.igem.org/mediawiki/igem.org/8/88/Logoigemklein.png" border="0" width="100" height="100"></a></div>
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| - | <img src="https://static.igem.org/mediawiki/igem.org/c/c6/Receptor_header.jpg" align="middle" width="100%">
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| - | <div id="contentbox" style="text-align:justify;">
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| - | <h2>Yeast for dummies</h2>
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| - | <p>Since this is the first time that TU Delfts iGEM team is working with yeast, we faced a lot of small yeast-related 'challenges'. With this page we want to inform you about the basics and the pitfalls of working with yeast. </p>
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| - | <h3>Auxotrophy</h3>
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| - | <h3>Shuttle vectors</h3>
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| - | <p> Today, most commonly encountered S. cerevisiae shuttle vectors belong to one of three classes:<br/>
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| - | - Integrating plasmids YIp<br/>
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| - | - Centromere plasmids YCp<br/>
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| - | - Yeast episomal plasmids YEp<br/>
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| - | We choose to work with the pRS vectors because.<br/>
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| - | The name pRS415 gives an indication on the presence of a CEN/ARS replication origin. 0 means yeast
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| - | integrative plasmid, 1 means that it also can be used to maintain the plasmid in circular
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| - | form.
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| - | pRS415 Gives an indication of the auxotrophic marker used.
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| - | pRS415 Version number… not really different.<br/>
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| - | <h3>Chromosomal integration</h3>
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| - | <p>We encountered a lot of problems with plasmids. Because we wanted our constructs to be universal
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| - | (with the idea to make it suitable for ‘fast checking’) we tried maintaining a plasmid. As it turned
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| - | out, yeast cells are not eager to maintain a plasmid and with our construct we suspect homologous
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| - | recombination occurred. After transformation, a PCR on the transformed plasmid, obtained by
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| - | isolation, showed two bands instead of the suspected single band, one being ! Integration of the
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| - | plasmid is therefore advised! Checking of this can be quite gruesome optimizing the necessary PCR
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| - | reactions on your transformed yeast colonies. Chromosomal isolation can therefore improve the
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| - | steps. </p><br/>
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| - | <h3>Knock-out strains</h3>
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| - | <p>European iGEM teams have the advantage to have Euroscarf available (http://web.uni-
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| - | frankfurt.de/fb15/mikro/euroscarf) to order strains with knocked out ORFs. The typical
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| - | nomenclature is also explained here (http://web.uni-frankfurt.de/fb15/mikro/euroscarf/
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| - | stra_des.html).</p><br/>
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| - | </div>
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| - | <img src="https://static.igem.org/mediawiki/igem.org/3/37/Footer_2.jpg" align="middle" width="690">
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| - | <a href='https://2012.igem.org/Main_Page' target="_blank"><div id='logo_igem2'></div><a/></body></html>
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