Difference between revisions of "Team:Paris Bettencourt/Project/VitaminB2"
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The chromosome integration plasmid pKV6 was first altered to allow the insertion of the four genes by Golden Gate Assembly. This new plasmid was called p15.01.<br> | The chromosome integration plasmid pKV6 was first altered to allow the insertion of the four genes by Golden Gate Assembly. This new plasmid was called p15.01.<br> | ||
Then, the four genes were successfully inserted in p15.01 and cloned in <i>E. coli</i>.<br> | Then, the four genes were successfully inserted in p15.01 and cloned in <i>E. coli</i>.<br> | ||
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<img width="350px" src="https://static.igem.org/mediawiki/2015/6/61/PkV6_Map.jpg"/><br> | <img width="350px" src="https://static.igem.org/mediawiki/2015/6/61/PkV6_Map.jpg"/><br> | ||
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<div class="column-right" align="justify"> | <div class="column-right" align="justify"> | ||
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After cloning, cells began to release riboflavin in their surrounding media, suggesting that the expression system was functioning in E. coli as well. <br> | After cloning, cells began to release riboflavin in their surrounding media, suggesting that the expression system was functioning in E. coli as well. <br> | ||
Riboflavin concentration corresponding to the fluorescence (at 565nm) of the supernatant was estimated on a standard curve.<br> | Riboflavin concentration corresponding to the fluorescence (at 565nm) of the supernatant was estimated on a standard curve.<br> | ||
− | <img width=" | + | <img width="450px" src="https://static.igem.org/mediawiki/2015/6/6f/Riboflavin_standard_curve.png"><br> |
<b>Figure 5: </b><i>Standard curve of riboflavin fluorescence</i><br><br> | <b>Figure 5: </b><i>Standard curve of riboflavin fluorescence</i><br><br> | ||
From the standard curve, we derived a linear equation mapping riboflavin concentration to fluorescence (fluorescence = 16344*x + 418).<br> | From the standard curve, we derived a linear equation mapping riboflavin concentration to fluorescence (fluorescence = 16344*x + 418).<br> | ||
− | <img width=" | + | <img width="500px" src="https://static.igem.org/mediawiki/2015/8/80/PB_overproduction_riboflavin.png"><br> |
<b>Figure 6: </b><i>Over-production of riboflavin by E.coli </i><br><br> | <b>Figure 6: </b><i>Over-production of riboflavin by E.coli </i><br><br> | ||
<i>E. coli</i> strains over-produce riboflavin up to 6µg/mL of media.<br><br> | <i>E. coli</i> strains over-produce riboflavin up to 6µg/mL of media.<br><br> | ||
− | <img width=" | + | <img width="300px" src="https://static.igem.org/mediawiki/parts/d/d2/B2_plate_riboflavin_diffusing_m9.png"><br> |
− | <b>Figure 7: </b><i> | + | <b>Figure 7: </b><i>E. coli strain diffusing riboflavin in plate</i><br><br> |
+ | <img width="400px" src="https://static.igem.org/mediawiki/2015/9/92/PB_projetB2_pic7.jpg"><br> | ||
+ | <b>Figure 8: </b><i>Riboflavin revelation by fluorescence</i><br><br> | ||
We characterized the functioning of promoter p48 in <i>E. coli</i> and BioBrick it (<a href="http://parts.igem.org/Part:BBa_K1678004">BBa_K1678004</a>).<br> | We characterized the functioning of promoter p48 in <i>E. coli</i> and BioBrick it (<a href="http://parts.igem.org/Part:BBa_K1678004">BBa_K1678004</a>).<br> | ||
Activity of p48 was shown by the expression of the fluorescent protein mCerulean.<br> | Activity of p48 was shown by the expression of the fluorescent protein mCerulean.<br> | ||
− | <img width=" | + | <img width="400px" src="https://static.igem.org/mediawiki/parts/b/bb/BB_fluorescence_chart_BB_04.png"><br> |
− | <b>Figure | + | <b>Figure 9: </b><i>Fluorescence of mCerulean promoted by p48</i><br><br> |
</div> | </div> |
Latest revision as of 23:45, 19 November 2015