Team:TU-Eindhoven/Notebook/Week5

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<h3>Our lab work</h3>
<h3>Our lab work</h3>
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The transformation last time did not yield any colonies, neither did the transformation with the pYES/LacZ vector as a positive control. Therefore, the conclusion: <span class= "red">either the transformation isn't working, the plasmids aren't in order or it's the yeast strain that fails to grow</span>. We are going to transform Mid1 and the GECOs in another strain (EBY100) as a positive control. So we started over again... We made the plates, we did all the transformations of the three GECOs and one part of the channels. We couldn’t do the transformation of the other part of the channels, because we used a mix which included Leu for the media, which is the selection marker of this part. While we were waiting on the transformations in yeast, we redid the transformation of the pYES3 vector with the G-GECO into the Nova Blue E. coli. Furthermore, we tried to clean the protein solution by removing the Ca<sup>2+</sup> ions. But since someone (we still don't know who did it) added some NaCl, we had to start over. The NaCl stabilized the protein, so the Ca<sup> 2+</sup> couldn't be released. At the end of the week, we got the results of the transformation:<span class= "red"> the transformation succeeded</span>! We put the colonies in 20ml tubes to multiply. We also ran a gel to check the colonies of the Nova Blue E. coli and concluded that this transformation worked as well!
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The transformation last time did not yield any colonies, neither did the transformation with the pYES/LacZ vector as a positive control. Therefore, the conclusion: <span class= "red">either the transformation isn't working, the plasmids aren't in order or it's the yeast strain that fails to grow</span>. We are going to transform MID1 and the GECOs in another strain (EBY100) as a positive control. So we started over again... We made the plates, we did all the transformations of the three GECOs and one part of the channels. We couldn’t do the transformation of the other part of the channels, because we used a mix which included Leu for the media, which is the selection marker of this part. While we were waiting on the transformations in yeast, we redid the transformation of the pYES3 vector with the G-GECO into the Nova Blue E. coli. Furthermore, we tried to clean the protein solution by removing the Ca<sup>2+</sup> ions. But since someone (we still don't know who did it) added some NaCl, we had to start over. The NaCl stabilized the protein, so the Ca<sup> 2+</sup> couldn't be released. At the end of the week, we got the results of the transformation:<span class= "red"> the transformation succeeded</span>! We put the colonies in 20 ml tubes to multiply. We also ran a gel to check the colonies of the Nova Blue E. coli and concluded that this transformation worked as well!

Revision as of 01:13, 27 September 2012