Team:CIDEB-UANL Mexico/project aroma

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<p align="justify">Since the beginning of iGEM project, the use of fluorescent reporters has been used in each one of the proposed projects in previous years, trying to test the theoretical presence of other proteins in <i>E. coli.</i> For our iGEM 2014 project, this module proposed to promote the usage of aroma reporters, instead of fluorescent ones.</p>
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<p><b><h2>Description</h2></b></p>
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<p>In the module, WinterGreen is the most important part. Normally, it catalyzes the conversion of salicylic acid into methyl salicylate. The protein is excreted by the bacteria and when salicylic acid is added to the medium, a chemical reaction takes place and produces methyl salicylate, which is responsible for the wintergreen odor.</p>
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      <h2>Aroma Module</h2>
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<p>But in this module, it will be regulated by temperature with the use of the RNA thermometer. When adding salicylic acid to the bacteria in a 32° Celsius environment, the production of the WinterGreen protein will begin.</p><br>
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<p>Since the beginning of iGEM project, the use of fluorescent reporters has been used in each one of the proposed projects in previous years, trying to test the theoretical presence of other proteins in E. coli. For our iGEM 2014 project, this module proposed to promote the usage of aroma reporters, instead of fluorescent ones.</p>
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<td style="padding-left:12px;"><img width=200 height=150 src="https://static.igem.org/mediawiki/2014hs/0/06/AromaHoja.png"/></td>
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<p><b>How is the Aroma module composed?</b></p>
 
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<p>This gene is composed by the following parts (see <b>figure 1A</b>): (1) a constitutive promoter, (2) a RNA thermometer, also called ribo-switch; used to regulate the WinterGreen-odor protein production through temperature, (3) a Wintergreen-odor enzyme generator, used to allow the production of methyl salicylate, induced by salicylic acid, and (4)  a terminator. All of these parts are ligated by an 8-bp scar (TACTAGAG).</p>
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<center><img width=350 height=250 src="https://static.igem.org/mediawiki/2014hs/6/6e/AromaFigure1Bb.png" align=center hspace=12/></center>
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<center><p><b>Figure 1.</b> Production of Wintergreen Odor</p></center>
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<center><p><img width=504 height=302 src="https://static.igem.org/mediawiki/2014hs/0/06/AromaFigure1A.png"
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<p><b><h2>Device</h2></b></p>
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<p>Originally, WinterGreen was thought to be the reporter of the Capture module, but in order to prove the function of this gene and also the one in charge of the capture of sodium ions (NhaS), we decided to separate the full device into the actual modules of Capture and Aroma, as seen in the figure below.</p>
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<p><b>Figure 1A</b> Aroma Module</p></center>
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<p><img width=450 height=250 src="https://static.igem.org/mediawiki/2014hs/3/32/TestCaptureAndAroma.jpg" align=center hspace=12></p>
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<p><b>Figure 2.</b> The device originally thought, and the 2 modules derived from it.</p></center><br>
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<p>Different to fluorescent reporters, this module was made in order to perform as an aroma reporter and also to test the correct function of the bacteria, for its future usage as a new reporter and functional part (CDS). It is desired to use this part in the project to replace the red fluorescent protein (RFP) in the Capture module. But it was preferable to test it apart to demonstrate its effectiveness. Similarly, this piece is also helpful for Union module, because when performing the filtration by silica, WinterGreen can demonstrate the presence of bacteria in the beads. The team added the RNA thermometer for regulating the production of the aroma in the project.</p>
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<p>This device is composed by the following parts (see <b>figure 3</b>): a constitutive promoter (1), a RNA thermometer (3); used to regulate the WinterGreen-odor protein production through temperature, a Wintergreen-odor enzyme generator, used to allow the production of methyl salicylate, induced by salicylic acid (2); and a terminator (4). All of these parts are ligated by an 8-bp scar (TACTAGAG).</p>
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<p><b>How does it work?</b></p>
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<center><img width=400 height=170 src="https://static.igem.org/mediawiki/2014hs/5/56/AromaFigure1Aa.png" align=center hspace=12/>
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<p><b>Figure 3.</b> Aroma Module</p></center><br>
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<center><p><img width=504 height=302 src="https://static.igem.org/mediawiki/2014hs/b/bb/AromaFigure1B.png"
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<p>These 4 genetic parts form the Aroma device of the project. The full device's length is 1,251bp (including restriction sites).</p>
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align=center hspace=12 alt="IMG_0317"></p>
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<p><b>Figure 1B</b> Production of Wintergreen Odor</p></center>
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<p><b><h2>Parts of the module</h2></b></p>
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<br><center><div><table class=MsoTable15Grid7ColorfulAccent3 border=1 cellspacing=0 cellpadding=0 width=625 style='width:468.55pt;border-collapse:collapse; border:none;mso-border-alt:solid #C2D69B .5pt;mso-border-themecolor:accent3; mso-border-themetint:153;mso-yfti-tbllook:1184;mso-padding-alt:0cm 5.4pt 0cm 5.4pt'> <tr style='mso-yfti-irow:-1;mso-yfti-firstrow:yes;mso-yfti-lastfirstrow:yes; height:15.0pt'> <td width=88 nowrap valign=top style='width:66.1pt;border:none;border-bottom: solid #C2D69B 1.0pt;mso-border-bottom-themecolor:accent3;mso-border-bottom-themetint: 153;mso-border-bottom-alt:solid #C2D69B .5pt;mso-border-bottom-themecolor: accent3;mso-border-bottom-themetint:153;background:white;mso-background-themecolor: background1;padding:0cm 5.4pt 0cm 5.4pt;height:15.0pt'> <p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal;mso-yfti-cnfc:517'><b><span lang=ES-MX style='font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:"Times New Roman";mso-fareast-language:ES-MX;mso-bidi-font-style: italic'>IMAGE</span></b><span lang=ES-MX style='font-family:Oxygen; mso-fareast-font-family:"Times New Roman";mso-bidi-font-family:"Times New Roman"; mso-fareast-language:ES-MX;mso-bidi-font-style:italic'><o:p></o:p></span></p> </td> <td width=119 valign=top style='width:89.0pt;border:none;border-bottom:solid #C2D69B 1.0pt; mso-border-bottom-themecolor:accent3;mso-border-bottom-themetint:153; mso-border-bottom-alt:solid #C2D69B .5pt;mso-border-bottom-themecolor:accent3; mso-border-bottom-themetint:153;background:white;mso-background-themecolor: background1;padding:0cm 5.4pt 0cm 5.4pt;height:15.0pt'> <p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal;mso-yfti-cnfc:1'><b><span lang=ES-MX style='font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:"Times New Roman";mso-fareast-language:ES-MX'>CODE</span></b><span lang=ES-MX style='font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:"Times New Roman";mso-fareast-language:ES-MX'><o:p></o:p></span></p> </td> <td width=418 nowrap valign=top style='width:313.45pt;border:none;border-bottom: solid #C2D69B 1.0pt;mso-border-bottom-themecolor:accent3;mso-border-bottom-themetint: 153;mso-border-bottom-alt:solid #C2D69B .5pt;mso-border-bottom-themecolor: accent3;mso-border-bottom-themetint:153;background:white;mso-background-themecolor: background1;padding:0cm 5.4pt 0cm 5.4pt;height:15.0pt'> <p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal;mso-yfti-cnfc:1'><b><span lang=ES-MX style='font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:"Times New Roman";mso-fareast-language:ES-MX'>DESCRIPTION</span></b><span lang=ES-MX style='font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:"Times New Roman";mso-fareast-language:ES-MX'><o:p></o:p></span></p> </td> </tr> <tr style='mso-yfti-irow:0;height:83.15pt'> <td width=88 nowrap valign=top style='width:66.1pt;border:none;border-right: solid #C2D69B 1.0pt;mso-border-right-themecolor:accent3;mso-border-right-themetint: 153;mso-border-top-alt:solid #C2D69B .5pt;mso-border-top-themecolor:accent3; mso-border-top-themetint:153;mso-border-top-alt:solid #C2D69B .5pt; mso-border-top-themecolor:accent3;mso-border-top-themetint:153;mso-border-right-alt: solid #C2D69B .5pt;mso-border-right-themecolor:accent3;mso-border-right-themetint: 153;background:white;mso-background-themecolor:background1;padding:0cm 5.4pt 0cm 5.4pt; height:83.15pt'> <p class=MsoNormal align=right style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:right;line-height:normal;mso-yfti-cnfc:68'><!--[if gte vml 1]><v:shape id="_x0000_s1029" type="#_x0000_t75" style='position:absolute;left:0; text-align:left;margin-left:10.95pt;margin-top:3.2pt;width:66pt;height:48pt; z-index:251660800;visibility:visible;mso-wrap-style:square; mso-width-percent:0;mso-height-percent:0;mso-wrap-distance-left:9pt; mso-wrap-distance-top:0;mso-wrap-distance-right:9pt; mso-wrap-distance-bottom:0;mso-position-horizontal:absolute; mso-position-horizontal-relative:text;mso-position-vertical:absolute; mso-position-vertical-relative:text;mso-width-percent:0; mso-height-percent:0;mso-width-relative:page;mso-height-relative:page'> <v:imagedata src="TablasAromaYResistencia_archivos/image007.png" o:title=""/> <w:wrap type="square"/> </v:shape><![endif]--><![if !vml]><img width=88 height=64 src="https://static.igem.org/mediawiki/2014hs/4/43/PConsAroma.png" align=left hspace=12 v:shapes="_x0000_s1029"><![endif]></p> <p class=MsoNormal align=right style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:right;line-height:normal;mso-yfti-cnfc:68'><i><span lang=ES-MX style='font-family:Oxygen;color:#76923C;mso-themecolor:accent3;mso-themeshade: 191;mso-fareast-language:ES-MX;mso-no-proof:yes'><o:p>&nbsp;</o:p></span></i></p> </td> <td width=119 nowrap valign=top style='width:89.0pt;border-top:none; border-left:none;border-bottom:solid #C2D69B 1.0pt;mso-border-bottom-themecolor: accent3;mso-border-bottom-themetint:153;border-right:solid #C2D69B 1.0pt; mso-border-right-themecolor:accent3;mso-border-right-themetint:153; mso-border-top-alt:solid #C2D69B .5pt;mso-border-top-themecolor:accent3; mso-border-top-themetint:153;mso-border-left-alt:solid #C2D69B .5pt; mso-border-left-themecolor:accent3;mso-border-left-themetint:153;mso-border-alt: solid #C2D69B .5pt;mso-border-themecolor:accent3;mso-border-themetint:153; background:#EAF1DD;mso-background-themecolor:accent3;mso-background-themetint: 51;padding:0cm 5.4pt 0cm 5.4pt;height:83.15pt'> <p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal;mso-yfti-cnfc:64'><span lang=ES-MX style='font-size:12.0pt;mso-bidi-font-size:11.0pt;font-family:Oxygen; mso-fareast-font-family:"Times New Roman";mso-bidi-font-family:Arial; color:black;mso-fareast-language:ES-MX'><a href="http://parts.igem.org/Part:BBa_J23119">BBa_J23119A</a></span><span lang=ES-MX style='font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:Arial;color:black;mso-fareast-language:ES-MX'><o:p></o:p></span></p> <p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal;mso-yfti-cnfc:64'>&nbsp;</p> </td> <td width=418 valign=top style='width:313.45pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;mso-border-bottom-themecolor:accent3; mso-border-bottom-themetint:153;border-right:solid #C2D69B 1.0pt;mso-border-right-themecolor: accent3;mso-border-right-themetint:153;mso-border-top-alt:solid #C2D69B .5pt; mso-border-top-themecolor:accent3;mso-border-top-themetint:153;mso-border-left-alt: solid #C2D69B .5pt;mso-border-left-themecolor:accent3;mso-border-left-themetint: 153;mso-border-alt:solid #C2D69B .5pt;mso-border-themecolor:accent3; mso-border-themetint:153;background:#EAF1DD;mso-background-themecolor:accent3; mso-background-themetint:51;padding:0cm 5.4pt 0cm 5.4pt;height:83.15pt'> <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;text-align: justify;line-height:normal;mso-yfti-cnfc:64'><span style='font-size:12.0pt; mso-bidi-font-size:11.0pt;font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:Arial;color:black;mso-ansi-language:EN-US;mso-fareast-language: ES-MX'>In the specific case of our aroma module, it will help the bacteria to continuously transcribe the <span class=SpellE>WinterGreen</span> gene in order to allow the bacteria to continuously produce the aroma. </span><span class=SpellE><span lang=ES-MX style='font-size:12.0pt;mso-bidi-font-size: 11.0pt;font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:Arial;color:black;mso-fareast-language:ES-MX'>This</span></span><span lang=ES-MX style='font-size:12.0pt;mso-bidi-font-size:11.0pt;font-family: Oxygen;mso-fareast-font-family:"Times New Roman";mso-bidi-font-family:Arial; color:black;mso-fareast-language:ES-MX'> <span class=SpellE>promoter</span> has a <span class=SpellE>length</span> of 35bp.<o:p></o:p></span></p> </td> </tr> <tr style='mso-yfti-irow:1;height:77.45pt'> <td width=88 nowrap valign=top style='width:66.1pt;border-top:solid #C2D69B 1.0pt; mso-border-top-themecolor:accent3;mso-border-top-themetint:153;border-left: none;border-bottom:none;border-right:solid #C2D69B 1.0pt;mso-border-right-themecolor: accent3;mso-border-right-themetint:153;mso-border-top-alt:solid #C2D69B .5pt; mso-border-top-themecolor:accent3;mso-border-top-themetint:153;mso-border-right-alt: solid #C2D69B .5pt;mso-border-right-themecolor:accent3;mso-border-right-themetint: 153;background:white;mso-background-themecolor:background1;padding:0cm 5.4pt 0cm 5.4pt; height:77.45pt'> <p class=MsoNormal align=right style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:right;line-height:normal;mso-yfti-cnfc:4'><!--[if gte vml 1]><v:shape id="_x0000_s1028" type="#_x0000_t75" style='position:absolute;left:0; text-align:left;margin-left:16.95pt;margin-top:4.25pt;width:57.75pt; height:44.25pt;z-index:251662848;visibility:visible;mso-wrap-style:square; mso-width-percent:0;mso-height-percent:0;mso-wrap-distance-left:9pt; mso-wrap-distance-top:0;mso-wrap-distance-right:9pt; mso-wrap-distance-bottom:0;mso-position-horizontal:absolute; mso-position-horizontal-relative:text;mso-position-vertical:absolute; mso-position-vertical-relative:text;mso-width-percent:0; mso-height-percent:0;mso-width-relative:page;mso-height-relative:page'> <v:imagedata src="TablasAromaYResistencia_archivos/image009.png" o:title=""/> <w:wrap type="square"/> </v:shape><![endif]--><![if !vml]><img width=77 height=59 src="https://static.igem.org/mediawiki/2014hs/b/b0/RNAthermAroma.png" align=left hspace=12 v:shapes="_x0000_s1028"><![endif]><i><span lang=ES-MX style='font-family: Oxygen;color:#76923C;mso-themecolor:accent3;mso-themeshade:191;mso-fareast-language: ES-MX;mso-no-proof:yes'><o:p></o:p></span></i></p></td> <td width=119 nowrap valign=top style='width:89.0pt;border-top:none; border-left:none;border-bottom:solid #C2D69B 1.0pt;mso-border-bottom-themecolor: accent3;mso-border-bottom-themetint:153;border-right:solid #C2D69B 1.0pt; mso-border-right-themecolor:accent3;mso-border-right-themetint:153; mso-border-top-alt:solid #C2D69B .5pt;mso-border-top-themecolor:accent3; mso-border-top-themetint:153;mso-border-left-alt:solid #C2D69B .5pt; mso-border-left-themecolor:accent3;mso-border-left-themetint:153;mso-border-alt: solid #C2D69B .5pt;mso-border-themecolor:accent3;mso-border-themetint:153; padding:0cm 5.4pt 0cm 5.4pt;height:77.45pt'> <p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal'><span lang=ES-MX style='font-size:12.0pt; mso-bidi-font-size:11.0pt;font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:Arial;color:black;mso-fareast-language:ES-MX'><a href="http://parts.igem.org/Part:BBa_K115017">BBa_K115017</a><o:p></o:p></span></p></td> <td width=418 valign=top style='width:313.45pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;mso-border-bottom-themecolor:accent3; mso-border-bottom-themetint:153;border-right:solid #C2D69B 1.0pt;mso-border-right-themecolor: accent3;mso-border-right-themetint:153;mso-border-top-alt:solid #C2D69B .5pt; mso-border-top-themecolor:accent3;mso-border-top-themetint:153;mso-border-left-alt: solid #C2D69B .5pt;mso-border-left-themecolor:accent3;mso-border-left-themetint: 153;mso-border-alt:solid #C2D69B .5pt;mso-border-themecolor:accent3; mso-border-themetint:153;padding:0cm 5.4pt 0cm 5.4pt;height:77.45pt'> <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;text-align: justify;line-height:normal'><span style='font-size:12.0pt;mso-bidi-font-size: 11.0pt;font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:Arial;color:black;mso-ansi-language:EN-US;mso-fareast-language: ES-MX'>A RNA thermometer, used for temperature post-transcriptional regulation (thermo sensor), and is designed to initiate transcription around 32°C.<o:p></o:p></span></p> </td> </tr> <tr style='mso-yfti-irow:2;height:190.6pt'> <td width=88 nowrap valign=top style='width:66.1pt;border-top:solid #C2D69B 1.0pt; mso-border-top-themecolor:accent3;mso-border-top-themetint:153;border-left: none;border-bottom:none;border-right:solid #C2D69B 1.0pt;mso-border-right-themecolor: accent3;mso-border-right-themetint:153;mso-border-top-alt:solid #C2D69B .5pt; mso-border-top-themecolor:accent3;mso-border-top-themetint:153;mso-border-right-alt: solid #C2D69B .5pt;mso-border-right-themecolor:accent3;mso-border-right-themetint: 153;background:white;mso-background-themecolor:background1;padding:0cm 5.4pt 0cm 5.4pt; height:190.6pt'> <p class=MsoNormal align=right style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:right;line-height:normal;mso-yfti-cnfc:68'><!--[if gte vml 1]><v:shape id="_x0000_s1027" type="#_x0000_t75" style='position:absolute;left:0; text-align:left;margin-left:13.95pt;margin-top:3.2pt;width:63pt;height:51.75pt; z-index:251664896;visibility:visible;mso-wrap-style:square; mso-width-percent:0;mso-height-percent:0;mso-wrap-distance-left:9pt; mso-wrap-distance-top:0;mso-wrap-distance-right:9pt; mso-wrap-distance-bottom:0;mso-position-horizontal:absolute; mso-position-horizontal-relative:text;mso-position-vertical:absolute; mso-position-vertical-relative:text;mso-width-percent:0; mso-height-percent:0;mso-width-relative:page;mso-height-relative:page'> <v:imagedata src="TablasAromaYResistencia_archivos/image011.png" o:title=""/> <w:wrap type="square"/> </v:shape><![endif]--><![if !vml]><img width=84 height=69 src="https://static.igem.org/mediawiki/2014hs/b/b1/WinterGreenAroma.png" align=left hspace=12 v:shapes="_x0000_s1027"><![endif]><i><span style='font-size:3.0pt;font-family: Oxygen;color:#76923C;mso-themecolor:accent3;mso-themeshade:191;mso-ansi-language: EN-US;mso-fareast-language:ES-MX;mso-no-proof:yes'><o:p></o:p></span></i></p></td> <td width=119 nowrap valign=top style='width:89.0pt;border-top:none; border-left:none;border-bottom:solid #C2D69B 1.0pt;mso-border-bottom-themecolor: accent3;mso-border-bottom-themetint:153;border-right:solid #C2D69B 1.0pt; mso-border-right-themecolor:accent3;mso-border-right-themetint:153; mso-border-top-alt:solid #C2D69B .5pt;mso-border-top-themecolor:accent3; mso-border-top-themetint:153;mso-border-left-alt:solid #C2D69B .5pt; mso-border-left-themecolor:accent3;mso-border-left-themetint:153;mso-border-alt: solid #C2D69B .5pt;mso-border-themecolor:accent3;mso-border-themetint:153; background:#EAF1DD;mso-background-themecolor:accent3;mso-background-themetint: 51;padding:0cm 5.4pt 0cm 5.4pt;height:190.6pt'> <p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal;mso-yfti-cnfc:64'><span style='font-size:12.0pt;mso-bidi-font-size:11.0pt;font-family:Oxygen; mso-fareast-font-family:"Times New Roman";mso-bidi-font-family:Arial; mso-ansi-language:EN-US;mso-fareast-language:ES-MX'><a href="http://parts.igem.org/Part:BBa_K1255001:Design">BBa_K1255001</a><o:p></o:p></span></p> <p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal;mso-yfti-cnfc:64'>&nbsp;</p> </td> <td width=418 valign=top style='width:313.45pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;mso-border-bottom-themecolor:accent3; mso-border-bottom-themetint:153;border-right:solid #C2D69B 1.0pt;mso-border-right-themecolor: accent3;mso-border-right-themetint:153;mso-border-top-alt:solid #C2D69B .5pt; mso-border-top-themecolor:accent3;mso-border-top-themetint:153;mso-border-left-alt: solid #C2D69B .5pt;mso-border-left-themecolor:accent3;mso-border-left-themetint: 153;mso-border-alt:solid #C2D69B .5pt;mso-border-themecolor:accent3; mso-border-themetint:153;background:#EAF1DD;mso-background-themecolor:accent3; mso-background-themetint:51;padding:0cm 5.4pt 0cm 5.4pt;height:190.6pt'> <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;text-align: justify;line-height:normal;mso-yfti-cnfc:64'><span style='font-size:12.0pt; mso-bidi-font-size:11.0pt;font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:Arial;color:black;mso-ansi-language:EN-US;mso-fareast-language: ES-MX'>Produces a <span class=SpellE>transferase</span> to convert salicylic acid into methyl salicylate (<span class=SpellE>WinterGreen</span> odor). The wintergreen odor generator requires of 2mM of salicylic acid to produce methyl salicylate. BSMT1 (<a href="http://parts.igem.org/Part:BBa_J45004">BBa_J45004</a>), <span class=SpellE>WinterGreen</span> Odor Generator original name, was created by <a href="https://2006.igem.org/wiki/index.php/MIT_2006">MIT 2006</a>. This year, the team optimized the sequence for <i>Escherichia coli</i>. </span><span class=SpellE><span lang=ES-MX style='font-size:12.0pt; mso-bidi-font-size:11.0pt;font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:Arial;color:black;mso-fareast-language:ES-MX'>The</span></span><span lang=ES-MX style='font-size:12.0pt;mso-bidi-font-size:11.0pt;font-family: Oxygen;mso-fareast-font-family:"Times New Roman";mso-bidi-font-family:Arial; color:black;mso-fareast-language:ES-MX'> new <span class=SpellE>biobrick</span> has a <span class=SpellE>length</span> of 1,074bp.<o:p></o:p></span></p> </td> </tr> <tr style='mso-yfti-irow:3;mso-yfti-lastrow:yes;height:70.7pt'> <td width=88 nowrap valign=top style='width:66.1pt;border-top:solid #C2D69B 1.0pt; mso-border-top-themecolor:accent3;mso-border-top-themetint:153;border-left: none;border-bottom:none;border-right:solid #C2D69B 1.0pt;mso-border-right-themecolor: accent3;mso-border-right-themetint:153;mso-border-top-alt:solid #C2D69B .5pt; mso-border-top-themecolor:accent3;mso-border-top-themetint:153;mso-border-right-alt: solid #C2D69B .5pt;mso-border-right-themecolor:accent3;mso-border-right-themetint: 153;background:white;mso-background-themecolor:background1;padding:0cm 5.4pt 0cm 5.4pt; height:70.7pt'> <p class=MsoNormal align=right style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:right;line-height:normal;mso-yfti-cnfc:4'><!--[if gte vml 1]><v:shape id="_x0000_s1026" type="#_x0000_t75" style='position:absolute;left:0; text-align:left;margin-left:18pt;margin-top:1.85pt;width:59.1pt;height:49.4pt; z-index:251666944;visibility:visible;mso-wrap-style:square; mso-width-percent:0;mso-height-percent:0;mso-wrap-distance-left:9pt; mso-wrap-distance-top:0;mso-wrap-distance-right:9pt; mso-wrap-distance-bottom:0;mso-position-horizontal:absolute; mso-position-horizontal-relative:text;mso-position-vertical:absolute; mso-position-vertical-relative:text;mso-width-percent:0; mso-height-percent:0;mso-width-relative:page;mso-height-relative:page'> <v:imagedata src="TablasAromaYResistencia_archivos/image013.png" o:title=""/> <w:wrap type="square"/> </v:shape><![endif]--><![if !vml]><img width=79 height=66 src="https://static.igem.org/mediawiki/2014hs/6/62/TerminatorAroma.png" align=left hspace=12 v:shapes="_x0000_s1026"><![endif]></p> </td> <td width=119 nowrap valign=top style='width:89.0pt;border-top:none; border-left:none;border-bottom:solid #C2D69B 1.0pt;mso-border-bottom-themecolor: accent3;mso-border-bottom-themetint:153;border-right:solid #C2D69B 1.0pt; mso-border-right-themecolor:accent3;mso-border-right-themetint:153; mso-border-top-alt:solid #C2D69B .5pt;mso-border-top-themecolor:accent3; mso-border-top-themetint:153;mso-border-left-alt:solid #C2D69B .5pt; mso-border-left-themecolor:accent3;mso-border-left-themetint:153;mso-border-alt: solid #C2D69B .5pt;mso-border-themecolor:accent3;mso-border-themetint:153; padding:0cm 5.4pt 0cm 5.4pt;height:70.7pt'> <p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal'><span lang=ES-MX style='font-size:12.0pt; mso-bidi-font-size:11.0pt;font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:Arial;mso-fareast-language:ES-MX'><a href="http://parts.igem.org/Part:BBa_B1002">BBa_B1002</a><o:p></o:p></span></p> <p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal'>&nbsp;</p> </td> <td width=418 valign=top style='width:313.45pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;mso-border-bottom-themecolor:accent3; mso-border-bottom-themetint:153;border-right:solid #C2D69B 1.0pt;mso-border-right-themecolor: accent3;mso-border-right-themetint:153;mso-border-top-alt:solid #C2D69B .5pt; mso-border-top-themecolor:accent3;mso-border-top-themetint:153;mso-border-left-alt: solid #C2D69B .5pt;mso-border-left-themecolor:accent3;mso-border-left-themetint: 153;mso-border-alt:solid #C2D69B .5pt;mso-border-themecolor:accent3; mso-border-themetint:153;padding:0cm 5.4pt 0cm 5.4pt;height:70.7pt'> <p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;text-align: justify;line-height:normal'><span style='font-size:12.0pt;mso-bidi-font-size: 11.0pt;font-family:Oxygen;mso-fareast-font-family:"Times New Roman"; mso-bidi-font-family:Arial;color:black;mso-ansi-language:EN-US;mso-fareast-language: ES-MX'>Part made of 6bp, responsible for transcription stop. The terminator stops the production of methyl salicylate. <o:p></o:p></span></p> </td> </tr></table></div></center><br>
-
<p>Besides, this module has a constitutive promoter which will be regulated by temperature with the use of the RNA thermometer. When adding salicylic acid to the bacteria in a 32° Celsius environment, the production of the WinterGreen protein will begin.</p>
+
<p><b><h2>Justification</h2></b></p>
 +
<p>Different to fluorescent reporters, this module was made in order to (in the future) perform as an aroma reporter and also to test the correct function of the bacteria, for its future usage as a new reporter and functional part (CDS). It is desired to use this part in the project to replace the red fluorescent protein (RFP) in the Capture module. But it was preferable to test it apart to demonstrate its effectiveness. Similarly, this piece is also helpful when the biofilter is assembled, because when performing the filtration by silica, WinterGreen can demonstrate the presence of bacteria in the beads. </p>
-
<p><b>Parts of the module</b></p>
+
<p>The team added the RNA thermometer in the device for regulating the production of the aroma. Another reason for selecting the RNA thermometer as a regulator was to continue the CIDEB UANL 2013 work with it.</p>
-
<table width=100%>
+
<p><b><h2>Other teams that used RNA thermometer and BSMT1</h2></b></p>
-
<tr>
+
-
<td><img width=100 height=100 src="https://static.igem.org/mediawiki/2014hs/4/43/PConsAroma.png"/></td>
+
-
<td><p><b>Constitutive Promoter (BBa_J23119A)</b></p>
+
-
<p>A promoter refers to the part of DNA that initiates the gene transcription. A constitutive promoter refers to a promoter that is continuously working. In the specific case of our Aroma module, it will help the bacteria to continuously transcribe the WinterGreen (BSMT1) gene in order to allow the bacteria to continuously produce the aroma. This promoter has a length of 35bp.</p></td>
+
-
</tr>
+
-
<tr>
+
-
<td><img width=100 height=100 src="https://static.igem.org/mediawiki/2014hs/b/b0/RNAthermAroma.png"/></td>
+
-
<td><p><b>RNA Thermometer (BBa_K115017)</b></p>
+
-
<p>This RBS (Ribosome Binding Site) is classified as a RNA thermometer, used for temperature post-transcriptional regulation (thermo sensor), and is designed to initiate translation around 32°C. The thermo sensor is repressed when the temperature is below 32°C, it does not translate because the mRNA takes a secondary structure just as the image below and cannot enter the ribosome for translation; when the temperatures reaches a range between 32°C and 37°C, the thermo sensor is activated and the mRNA unfolds into a straight structure that now allows access to the ribosome to start the production of the protein. </p></td>
+
-
</tr>
+
-
</table>
+
-
<p>The following image (<b>Figure 2</b>) shows the secondary structure of the part after ligation to a protein coding part, as predicted by RNAfold. Notice that the 3' end, including the scar and the start codon, does not belong to the part. The blue region is the Shine Dalgarno (SD) sequence which is the ribosome binding site (TUDelft, 2008).</p>
+
<div align=center width="80%" >
 +
<table border=1 cellspacing=0 cellpadding=0 style='border-collapse:collapse;border:none;mso-border-alt:solid #C2D69B .5pt; mso-yfti-tbllook:1184;mso-padding-alt:0cm 5.4pt 0cm 5.4pt;mso-border-insideh: .5pt solid #C2D69B;mso-border-insidev:.5pt solid #C2D69B'>  
 +
<tr style='mso-yfti-irow:0;mso-yfti-firstrow:yes'>  
 +
<td width=170 valign=top style='width:127.35pt;border:solid #9BBB59 1.0pt; border-right:none;mso-border-top-alt:solid #9BBB59 .5pt;mso-border-left-alt: solid #9BBB59 .5pt;mso-border-bottom-alt:solid #9BBB59 .5pt;background:#9BBB59; padding:0cm 5.4pt 0cm 5.4pt'>  
-
<center><p><img width=504 height=302 src="https://static.igem.org/mediawiki/2014hs/8/8e/AromaFigure2.png"
+
<p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal'><b>
-
align=center hspace=12 alt="IMG_0317"></p>
+
-
<p><b>Figure 2.</b> RNA thermometer</p></center>
+
<span style='font-size:12.0pt; color:white;mso-ansi-language:EN-US'>Team<o:p></o:p>
 +
</span></b></p>  
 +
</td>
 +
<td width=170 valign=top style='width:127.35pt;border:solid #9BBB59 1.0pt; border-left:none;mso-border-top-alt:solid #9BBB59 .5pt;mso-border-bottom-alt: solid #9BBB59 .5pt;mso-border-right-alt:solid #9BBB59 .5pt;background:#9BBB59; padding:0cm 5.4pt 0cm 5.4pt'>  
-
<p>In the Aroma module, the thermo sensor will give us control over the production of the WinterGreen aroma. This RNA thermometer has a length of 83bp.</p>
+
<p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal'><b>
-
<table width=100%>
+
<span style='font-size:12.0pt; color:white;mso-ansi-language:EN-US'>Part Used<o:p></o:p>
-
<tr>
+
</span></b></p>  
-
<td><img width=100 height=100 src="https://static.igem.org/mediawiki/2014hs/b/b1/WinterGreenAroma.png"/></td>
+
</td>  
-
<td><p><b>WinterGreen Odor Generator (BSMT1) (BBa_J45004)</b></p>
+
<td width=387 valign=top style='width:290.6pt;border:solid #9BBB59 1.0pt; border-left:none;mso-border-top-alt:solid #9BBB59 .5pt;mso-border-bottom-alt: solid #9BBB59 .5pt;mso-border-right-alt:solid #9BBB59 .5pt;background:#9BBB59; padding:0cm 5.4pt 0cm 5.4pt'>  
-
<p>The CDS used in the Aroma module produces a transferase to convert salicylic acid into methyl salicylate, which main characteristic is the WinterGreen odor. The wintergreen odor generator requires of 2mM of salicylic acid to produce methyl salicylate. BSMT1 (WinterGreen Odor Generator original name) has a length of 1074bp.</p></td>
+
 
 +
<p class=MsoNormal align=center style='margin-bottom:0cm;margin-bottom:.0001pt; text-align:center;line-height:normal'><b>
 +
 
 +
<span style='font-size:12.0pt; color:white;mso-ansi-language:EN-US'>Use<o:p></o:p>
 +
</span></b></p>  
 +
</td>  
</tr>
</tr>
-
</table>
+
 +
<tr style='mso-yfti-irow:1;height:33.0pt'>
 +
<td width=170 valign=top style='width:127.35pt;border:solid #C2D69B 1.0pt; border-top:none;mso-border-top-alt:solid #C2D69B .5pt;mso-border-alt:solid #C2D69B .5pt; background:#EAF1DD;padding:0cm 5.4pt 0cm 5.4pt;height:33.0pt'>  
-
<p>This CDS works encoding SAM benzoic acid/salicylic acid carboxyl methyltransferase I, derived from BSMT1 from Petunia x hybrida. BSMT1 catalyzes the conversion of salicylic acid to methyl salicylate. Methyl benzoate has a floral smell and methyl salicylate has a wintergreen smell. (MIT, 2006).</p>
+
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'><b><u>
-
<center><p><img width=804 height=302 src="https://static.igem.org/mediawiki/2014hs/b/ba/AromaFigure3.png"
+
<span lang=FR style='font-size:12.0pt;color:#4F81BD;mso-ansi-language: FR'><a href="https://2008.igem.org/Team:TUDelft" target="_blank">TuDelft 2008</a>
-
align=center hspace=12 alt="IMG_0317"></p>
+
</span></u></b>
-
<p><b>Figure 3.</b> This chart shows the theoretical production of substrate by the induction of both benzoic and salicylic acids</p></center>
+
<span lang=FR style='font-size:6.0pt;mso-bidi-font-size:12.0pt;mso-ansi-language:FR; mso-bidi-font-weight:bold'>:
 +
</span><b>
-
<table width=100%>
+
<span lang=FR style='font-size:12.0pt; mso-ansi-language:FR'><o:p></o:p>
-
<tr>
+
</span></b></p>
-
<td><img width=100 height=100 src="https://static.igem.org/mediawiki/2014hs/6/62/TerminatorAroma.png"/></td>
+
</td>
-
<td><p><b>Terminator (BBa_B1002)</b></p>
+
<td width=170 valign=top style='width:127.35pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;border-right:solid #C2D69B 1.0pt; mso-border-top-alt:solid #C2D69B .5pt;mso-border-left-alt:solid #C2D69B .5pt; mso-border-alt:solid #C2D69B .5pt;background:#EAF1DD;padding:0cm 5.4pt 0cm 5.4pt; height:33.0pt'>  
-
<p>Part made of 6bp, responsible for transcription stop. The terminator stops the production of methyl salicylate.</p></td>
+
 
 +
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
 +
 
 +
<span style='font-size:12.0pt;mso-ansi-language:EN-US'>RNA themometer<o:p></o:p>
 +
</span></p>
 +
 
 +
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
 +
 
 +
<span style='font-size:12.0pt;mso-ansi-language:EN-US'><o:p>&nbsp;</o:p>
 +
</span></p>
 +
</td>  
 +
<td width=387 valign=top style='width:290.6pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;border-right:solid #C2D69B 1.0pt; mso-border-top-alt:solid #C2D69B .5pt;mso-border-left-alt:solid #C2D69B .5pt; mso-border-alt:solid #C2D69B .5pt;background:#EAF1DD;padding:0cm 5.4pt 0cm 5.4pt; height:33.0pt'>
 +
 
 +
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
 +
 
 +
<span style='font-size:12.0pt;mso-ansi-language:EN-US'>Temperature-sensing bacteria that changes color at different temperatures; as a temperature reporter system inlarge-scale fermentations, or as a temperature-inducible protein production system.<o:p></o:p>
 +
</span></p>
 +
 
 +
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
 +
 
 +
<span style='font-size:12.0pt;mso-ansi-language:EN-US'><o:p>&nbsp;</o:p>
 +
</span></p>
 +
</td>
 +
</tr>
 +
<tr style='mso-yfti-irow:2'>
 +
<td width=170 valign=top style='width:127.35pt;border:solid #C2D69B 1.0pt; border-top:none;mso-border-top-alt:solid #C2D69B .5pt;mso-border-alt:solid #C2D69B .5pt; padding:0cm 5.4pt 0cm 5.4pt'>
 +
 
 +
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'><b><u>
 +
 
 +
<span style='font-size:12.0pt;color:#4F81BD;mso-ansi-language: EN-US'><a href="https://2009.igem.org/Team:VictoriaBC" target="_blank">Victoria BC 2009</a>
 +
</span></u></b>
 +
 
 +
<span style='font-size:6.0pt;mso-bidi-font-size:12.0pt;mso-ansi-language:EN-US; mso-bidi-font-weight:bold'>:<b> <o:p></o:p></b>
 +
</span></p>
 +
 
 +
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'><b>
 +
 
 +
<span style='font-size:12.0pt;mso-ansi-language:EN-US'><o:p>&nbsp;</o:p>
 +
</span></b></p>
 +
</td>  
 +
<td width=170 valign=top style='width:127.35pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;border-right:solid #C2D69B 1.0pt; mso-border-top-alt:solid #C2D69B .5pt;mso-border-left-alt:solid #C2D69B .5pt; mso-border-alt:solid #C2D69B .5pt;padding:0cm 5.4pt 0cm 5.4pt'>  
 +
 
 +
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
 +
 
 +
<span style='font-size:12.0pt;mso-ansi-language:EN-US'>RNA themometer<o:p></o:p>
 +
</span></p>  
 +
 
 +
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
 +
 
 +
<span style='font-size:12.0pt;mso-ansi-language:EN-US'><o:p>&nbsp;</o:p>
 +
</span></p>
 +
</td>
 +
<td width=387 valign=top style='width:290.6pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;border-right:solid #C2D69B 1.0pt; mso-border-top-alt:solid #C2D69B .5pt;mso-border-left-alt:solid #C2D69B .5pt; mso-border-alt:solid #C2D69B .5pt;padding:0cm 5.4pt 0cm 5.4pt'>
 +
 
 +
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
 +
 
 +
<span style='font-size:12.0pt;mso-ansi-language:EN-US'>NAND logic gate using the ribo-key/ribo-lock system designed by <a href="https://2006.igem.org/wiki/index.php/Berkeley">Berkeley 2006</a> team , producing RFP except when the cells are grown in the presence of both arabinose and IPTG, also coupling fluorescent outputs with the ribo-thermometers made by TUDelft 2008 team.<o:p></o:p>
 +
</span></p>
 +
 
 +
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
 +
 
 +
<span style='font-size:12.0pt;mso-ansi-language:EN-US'><o:p>&nbsp;</o:p>
 +
</span></p>
 +
</td>  
</tr>
</tr>
-
</table>
+
<tr style='mso-yfti-irow:2'>
 +
<td width=170 valign=top style='width:127.35pt;border:solid #C2D69B 1.0pt; border-top:none;mso-border-top-alt:solid #C2D69B .5pt;mso-border-alt:solid #C2D69B .5pt; padding:0cm 5.4pt 0cm 5.4pt'>  
-
<p>These 4 genetic parts form the Aroma device of the project(see <b>Figure 4</b>).</p>
+
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'><b><u>
-
<center><p><img width=804 height=302 src="https://static.igem.org/mediawiki/2014hs/3/3a/AromaFigure4.png"
+
<span style='font-size:12.0pt;color:#4F81BD;mso-ansi-language: EN-US'><a href="https://2013hs.igem.org/Team:CIDEB-UANL_Mexico" target="_blank">CIDEB UANL 2013</a>
-
align=center hspace=12 alt="IMG_0317"></p>
+
</span></u></b>
-
<p><b>Figure 4</b> Aroma Device</p></center>
+
<span style='font-size:6.0pt;mso-bidi-font-size:12.0pt;mso-ansi-language:EN-US; mso-bidi-font-weight:bold'>:<b> <o:p></o:p></b>
 +
</span></p>  
-
<p><b>Other teams that use RNA thermometer and WinterGreen (BSMT1)</b></p>
+
<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'><b>
-
<p><i>RNA thermometer</i></p>
+
<span style='font-size:12.0pt;mso-ansi-language:EN-US'><o:p>&nbsp;</o:p>
 +
</span></b></p>
 +
</td>
 +
<td width=170 valign=top style='width:127.35pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;border-right:solid #C2D69B 1.0pt; mso-border-top-alt:solid #C2D69B .5pt;mso-border-left-alt:solid #C2D69B .5pt; mso-border-alt:solid #C2D69B .5pt;padding:0cm 5.4pt 0cm 5.4pt'>  
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<p class=MsoListParagraph style='margin-top:0cm;margin-right:0cm;margin-bottom:
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<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
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5.0pt;margin-left:21.3pt;'><![if !supportLists]><span
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style='font-size:12.0pt;font-family:Symbol;mso-fareast-font-family:Symbol;
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mso-bidi-font-family:Symbol'><span style='mso-list:Ignore'>·<span
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style='font:7.0pt "Times New Roman"'>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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</span></span></span><![endif]><span class=SpellE><b style='mso-bidi-font-weight:
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normal'><span>TUDelft</span></b></span><b
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style='mso-bidi-font-weight:normal'><span> 2008:</span></b><span> Temperature-sensing bacteria that changes
+
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color at different temperatures; as a temperature reporter system in
+
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large-scale fermentations, or as a temperature-inducible protein production
+
-
system. <o:p></o:p></span></p>
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<center><p><img width=704 height=302 src="https://static.igem.org/mediawiki/2014hs/e/e0/AromaFigure5.png"
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<span style='font-size:12.0pt;mso-ansi-language:EN-US'>RNA themometer<o:p></o:p>
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align=center hspace=12 alt="IMG_0317"></p>
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</span></p>  
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<p><b>Figure 5.</b> RNA Thermometer circuit, excerpted from TUDelft 2008 team</p></center>
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<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
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<span style='font-size:12.0pt;mso-ansi-language:EN-US'><o:p>&nbsp;</o:p>
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<td width=387 valign=top style='width:290.6pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;border-right:solid #C2D69B 1.0pt; mso-border-top-alt:solid #C2D69B .5pt;mso-border-left-alt:solid #C2D69B .5pt; mso-border-alt:solid #C2D69B .5pt;padding:0cm 5.4pt 0cm 5.4pt'>
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<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
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<span style='font-size:12.0pt;mso-ansi-language:EN-US'>Production of Vip3ca3, which acts as a pesticide protein, regulated by specific temperatures in order to avoid overproduction and it will show activity against target organisms Coleoptera and Lepidoptera..<o:p></o:p>
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<span style='font-size:12.0pt;mso-ansi-language:EN-US'><o:p>&nbsp;</o:p>
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<tr style='mso-yfti-irow:3;mso-yfti-lastrow:yes'>
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<td width=170 valign=top style='width:127.35pt;border:solid #C2D69B 1.0pt; border-top:none;mso-border-top-alt:solid #C2D69B .5pt;mso-border-alt:solid #C2D69B .5pt; background:#EAF1DD;padding:0cm 5.4pt 0cm 5.4pt'>
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<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'><b><u>
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<span lang=ES-MX style='font-size:12.0pt;color:#4F81BD'><a href="https://2006.igem.org/wiki/index.php/MIT_2006" target="_blank">MIT 2006</a>
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</span></u></b>
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<span lang=ES-MX style='font-size:7.0pt;mso-bidi-font-size:12.0pt;mso-bidi-font-weight: bold'>:<b> <o:p></o:p></b>
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<td width=170 valign=top style='width:127.35pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;border-right:solid #C2D69B 1.0pt; mso-border-top-alt:solid #C2D69B .5pt;mso-border-left-alt:solid #C2D69B .5pt; mso-border-alt:solid #C2D69B .5pt;background:#EAF1DD;padding:0cm 5.4pt 0cm 5.4pt'>
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<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
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<span style='font-size:12.0pt;mso-ansi-language:EN-US'>BSMT1<o:p></o:p>
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</span></p>
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</td>
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<td width=387 valign=top style='width:290.6pt;border-top:none;border-left: none;border-bottom:solid #C2D69B 1.0pt;border-right:solid #C2D69B 1.0pt; mso-border-top-alt:solid #C2D69B .5pt;mso-border-left-alt:solid #C2D69B .5pt; mso-border-alt:solid #C2D69B .5pt;background:#EAF1DD;padding:0cm 5.4pt 0cm 5.4pt'>
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<p class=MsoNormal style='margin-bottom:0cm;margin-bottom:.0001pt;line-height: normal'>
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 +
<span style='font-size:12.0pt;mso-ansi-language:EN-US'>This device produces methyl salicylate in the presence of salicylic acid. Methyl salicylate smells strongly of mint (wintergreen). Production of methyl salicylate was verified both by scent and by gas chromatography: E. coli with no WGD did not produce methyl salicylate when SA was added to the medium, while E. coli with the WGD did produce methyl salicylate when SA was added to the medium..<o:p></o:p>
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</span></p>
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</td>
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<p class=MsoListParagraph style='margin-top:0cm;margin-right:0cm;margin-bottom:
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<p><b><h2>Project Zoom in</h2></b></p>
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5.0pt;margin-left:21.3pt;'><![if !supportLists]><span
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<p><center><iframe width="600" height="500" src="//www.youtube.com/embed/fd9EQNCOr2k" frameborder="0" allowfullscreen></iframe></center></p>
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style='font-size:12.0pt;font-family:Symbol;mso-fareast-font-family:Symbol;
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<br>
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mso-bidi-font-family:Symbol'><span style='mso-list:Ignore'>·<span
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style='font:7.0pt "Times New Roman"'>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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</span></span></span><![endif]><span class=SpellE><b style='mso-bidi-font-weight:
+
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normal'><span> VictoriaBC</span></b></span><b
+
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style='mso-bidi-font-weight:normal'><span> 2009: </span></b><span>NAND logic gate using the ribo-key/ribo-lock system designed by the 2006 Berkeley team, producing RFP except when the cells are grown in the presence of both arabinose and IPTG, also coupling fluorescent outputs with the ribo-thermometers made by the 2008 TUDelft team. <o:p></o:p></span></p>
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<p class=MsoListParagraph style='margin-top:0cm;margin-right:0cm;margin-bottom:
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<p><b><h2>Bibliography/References</h2></b></p><font size="2">
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5.0pt;margin-left:21.3pt;'><![if !supportLists]><span
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style='font-size:12.0pt;font-family:Symbol;mso-fareast-font-family:Symbol;
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mso-bidi-font-family:Symbol'><span style='mso-list:Ignore'>·<span
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style='font:7.0pt "Times New Roman"'>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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</span></span></span><![endif]><span class=SpellE><b style='mso-bidi-font-weight:
+
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normal'><span>iGEM_CIDEB </span></b></span><b
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style='mso-bidi-font-weight:normal'><span>2013:</span></b><span> Production of Vip3ca3, which acts as a pesticide protein, regulated by specific temperatures in order to avoid overproduction and it will show activity against target organisms Coleoptera and Lepidoptera. <o:p></o:p></span></p>
+
-
<center><p><img width=704 height=302 src="https://static.igem.org/mediawiki/2014hs/3/3f/AromaFigure6.jpg "
+
<p>● Huang, H. (2006, August 30). <i>Part:BBa_B1002. </i> Retrieved August 30, 2014, from <a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_B1002"> http://parts.igem.org/wiki/index.php?title=Part:BBa_B1002</a>.</p>
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align=center hspace=12 alt="IMG_0317"></p>
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<p><b>Figure 6.</b> Circuit from iGEM CIDEB UANL 2013 team</p></center>
+
<p>● iGEM2006_Berkeley. (2006). <i>Part:BBa_J23100</i>. Retrieved August 30, 2014, from <a href="http://parts.igem.org/Part:BBa_J23100"> http://parts.igem.org/Part:BBa_J23100</a>.</p>
-
<p><i>WinterGreen (BSMT1)</i></p>
+
<p>● iGEM2006_MIT. (2006). <i>Part:BBa_J45004</i>. Retrieved August 30, 2014, from <a href="http://parts.igem.org/Part:BBa_J45004"> http://parts.igem.org/Part:BBa_J45004</a>. </p>
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<p class=MsoListParagraph style='margin-top:0cm;margin-right:0cm;margin-bottom:
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<p>● iGEM CIDEB Team. (2013). <i>iGEM CIDEB UANL 2013</i>. Retrieved on March 31th, 2014. <a href="https://2013hs.igem.org/Team:CIDEB-UANL_Mexico/Project"> https://2013hs.igem.org/Team:CIDEB-UANL_Mexico/Project</a>. </p>
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5.0pt;margin-left:21.3pt;'><![if !supportLists]><span
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style='font-size:12.0pt;font-family:Symbol;mso-fareast-font-family:Symbol;
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mso-bidi-font-family:Symbol'><span style='mso-list:Ignore'>·<span
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style='font:7.0pt "Times New Roman"'>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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</span></span></span><![endif]><span class=SpellE><b style='mso-bidi-font-weight:
+
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normal'><span>iGEM 2006_MIT:</span></b></span><span> This device produces methyl salicylate in the presence of salicylic acid. Methyl salicylate smells strongly of mint (wintergreen). Production of methyl salicylate was verified both by scent and by gas chromatography: E. coli with no WGD did not produce methyl salicylate when SA was added to the medium, while E. coli with the WGD did produce methyl salicylate when SA was added to the medium. <o:p></o:p></span></p>
+
-
<center><p><img width=604 height=302 src="https://static.igem.org/mediawiki/2014hs/9/98/AromaFigure7.jpg"
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<p>● iGEM08_TUDelft. (2008). <i>Part:BBa_K115017</i>. Retrieved August 30, 2014, from <a href="http://parts.igem.org/Part:BBa_K115017"> http://parts.igem.org/Part:BBa_K115017</a>. </p>
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align=center hspace=12 alt="IMG_0317"></p>
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<p><b>Figure 7.</b> Wintergreen odor enzyme (BSMT1) generator circuit</p></center>
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<p>● MIT IGEM Team. (2006). <i>MIT 2006</i>. Retrieved on March 31th, 2014, from: <a href="https://2006.igem.org/wiki/index.php/MIT_2006"> https://2006.igem.org/wiki/index.php/MIT_2006</a>. </p>
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<p><b>References</b></p>
+
<p>● TUDelft iGEM Team. (2008). <i>TUDelft 2008</i>. Retrieved on March 31th, 2014, from: <a href="https://2008.igem.org/Team:TUDelft"> https://2008.igem.org/Team:TUDelft</a>. </p>
-
<p>Zubieta, Chole et al. (2003). Structural Basis for Substrate Recognition in the Salicylic Acid Carboxyl Methyltransferase Family. Manuscript submitted for publication. Retrieved from www.plantcell.org; American Society of Plant Biologists.</p>
+
<p>● VictoriaBC. (2009). <i>VictoriaBC 2009</i>. Retrieved on March 31th, 2014, from: <a href="https://2009.igem.org/Team:VictoriaBC"> https://2009.igem.org/Team:VictoriaBC</a>. </p>
-
<p>Huang, H. (2006, August 30). Part:BBa_B1002. Retrieved August 30, 2014, from http://parts.igem.org/wiki/index.php?title=Part:BBa_B1002.</p>
+
-
<p>Part:BBa_J45004 (2006). Part:BBa_J45004. Retrieved August 30, 2014, from http://parts.igem.org/Part:BBa_J45004.</p>
+
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<p>Part:BBa_K115017 (2008). Part:BBa_K115017. Retrieved August 30, 2014, from http://parts.igem.org/Part:BBa_K115017.</p>
+
-
<p>Part:BBa_J23100 (2006). Part:BBa_J23100. Retrieved August 30, 2014, from http://parts.igem.org/Part:BBa_J23100.</p>
+
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<p>MIT IGEM Team. (2006). MIT 2006. Retrieved on March 31th, 2014. From: https://2006.igem.org/wiki/index.php/MIT_2006.</p>
+
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<p>TUDelft iGEM Team. (2008). TUDelft 2008. Retrieved on March 31th, 2014. From: https://2008.igem.org/Team:TUDelft.</p>
+
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<p>VictoriaBC. (2009). VictoriaBC 2009. Retrieved on March 31th, 2014. From: https://2009.igem.org/Team:VictoriaBC.</p>
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<p>iGEM CIDEB Team. (2013). iGEM CIDEB UANL 2013. Retrieved on March 31th, 2014. https://2013hs.igem.org/Team:CIDEB-UANL_Mexico/Project.</p>
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<div style="text-align: right;"><a href="https://2014hs.igem.org/Team:CIDEB-UANL_Mexico/project_aroma#"><font color="blue">Return to the Top</font></a></p></div>
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<p>● Zubieta, Chole et al. (2003). <i>Structural Basis for Substrate Recognition in the Salicylic Acid Carboxyl Methyltransferase Family</i>. Manuscript submitted for publication. Retrieved from <a href="www.plantcell.org"> www.plantcell.org</a>. </p></font>
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<br>
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<div style="text-align: right;"><a href="https://2014hs.igem.org/Team:CIDEB-UANL_Mexico/project_aroma#"><font color="blue">Return to the Top</font></a></p>
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Latest revision as of 03:27, 21 June 2014

Description Capture Module Aroma Module Resistance Module Union Module Parts
iGEM CIDEB 2014 - Project

Aroma Module


Since the beginning of iGEM project, the use of fluorescent reporters has been used in each one of the proposed projects in previous years, trying to test the theoretical presence of other proteins in E. coli. For our iGEM 2014 project, this module proposed to promote the usage of aroma reporters, instead of fluorescent ones.


Description

In the module, WinterGreen is the most important part. Normally, it catalyzes the conversion of salicylic acid into methyl salicylate. The protein is excreted by the bacteria and when salicylic acid is added to the medium, a chemical reaction takes place and produces methyl salicylate, which is responsible for the wintergreen odor.

But in this module, it will be regulated by temperature with the use of the RNA thermometer. When adding salicylic acid to the bacteria in a 32° Celsius environment, the production of the WinterGreen protein will begin.


Figure 1. Production of Wintergreen Odor

Device

Originally, WinterGreen was thought to be the reporter of the Capture module, but in order to prove the function of this gene and also the one in charge of the capture of sodium ions (NhaS), we decided to separate the full device into the actual modules of Capture and Aroma, as seen in the figure below.

Figure 2. The device originally thought, and the 2 modules derived from it.


This device is composed by the following parts (see figure 3): a constitutive promoter (1), a RNA thermometer (3); used to regulate the WinterGreen-odor protein production through temperature, a Wintergreen-odor enzyme generator, used to allow the production of methyl salicylate, induced by salicylic acid (2); and a terminator (4). All of these parts are ligated by an 8-bp scar (TACTAGAG).

Figure 3. Aroma Module


These 4 genetic parts form the Aroma device of the project. The full device's length is 1,251bp (including restriction sites).

Parts of the module


IMAGE

CODE

DESCRIPTION

 

BBa_J23119A

 

In the specific case of our aroma module, it will help the bacteria to continuously transcribe the WinterGreen gene in order to allow the bacteria to continuously produce the aroma. This promoter has a length of 35bp.

BBa_K115017

A RNA thermometer, used for temperature post-transcriptional regulation (thermo sensor), and is designed to initiate transcription around 32°C.

BBa_K1255001

 

Produces a transferase to convert salicylic acid into methyl salicylate (WinterGreen odor). The wintergreen odor generator requires of 2mM of salicylic acid to produce methyl salicylate. BSMT1 (BBa_J45004), WinterGreen Odor Generator original name, was created by MIT 2006. This year, the team optimized the sequence for Escherichia coli. The new biobrick has a length of 1,074bp.

BBa_B1002

 

Part made of 6bp, responsible for transcription stop. The terminator stops the production of methyl salicylate.


Justification

Different to fluorescent reporters, this module was made in order to (in the future) perform as an aroma reporter and also to test the correct function of the bacteria, for its future usage as a new reporter and functional part (CDS). It is desired to use this part in the project to replace the red fluorescent protein (RFP) in the Capture module. But it was preferable to test it apart to demonstrate its effectiveness. Similarly, this piece is also helpful when the biofilter is assembled, because when performing the filtration by silica, WinterGreen can demonstrate the presence of bacteria in the beads.

The team added the RNA thermometer in the device for regulating the production of the aroma. Another reason for selecting the RNA thermometer as a regulator was to continue the CIDEB UANL 2013 work with it.

Other teams that used RNA thermometer and BSMT1

Team

Part Used

Use

TuDelft 2008 :

RNA themometer

 

Temperature-sensing bacteria that changes color at different temperatures; as a temperature reporter system inlarge-scale fermentations, or as a temperature-inducible protein production system.

 

Victoria BC 2009 :

 

RNA themometer

 

NAND logic gate using the ribo-key/ribo-lock system designed by Berkeley 2006 team , producing RFP except when the cells are grown in the presence of both arabinose and IPTG, also coupling fluorescent outputs with the ribo-thermometers made by TUDelft 2008 team.

 

CIDEB UANL 2013 :

 

RNA themometer

 

Production of Vip3ca3, which acts as a pesticide protein, regulated by specific temperatures in order to avoid overproduction and it will show activity against target organisms Coleoptera and Lepidoptera..

 

MIT 2006 :

 

BSMT1

This device produces methyl salicylate in the presence of salicylic acid. Methyl salicylate smells strongly of mint (wintergreen). Production of methyl salicylate was verified both by scent and by gas chromatography: E. coli with no WGD did not produce methyl salicylate when SA was added to the medium, while E. coli with the WGD did produce methyl salicylate when SA was added to the medium..

Project Zoom in


Bibliography/References

● Huang, H. (2006, August 30). Part:BBa_B1002. Retrieved August 30, 2014, from http://parts.igem.org/wiki/index.php?title=Part:BBa_B1002.

● iGEM2006_Berkeley. (2006). Part:BBa_J23100. Retrieved August 30, 2014, from http://parts.igem.org/Part:BBa_J23100.

● iGEM2006_MIT. (2006). Part:BBa_J45004. Retrieved August 30, 2014, from http://parts.igem.org/Part:BBa_J45004.

● iGEM CIDEB Team. (2013). iGEM CIDEB UANL 2013. Retrieved on March 31th, 2014. https://2013hs.igem.org/Team:CIDEB-UANL_Mexico/Project.

● iGEM08_TUDelft. (2008). Part:BBa_K115017. Retrieved August 30, 2014, from http://parts.igem.org/Part:BBa_K115017.

● MIT IGEM Team. (2006). MIT 2006. Retrieved on March 31th, 2014, from: https://2006.igem.org/wiki/index.php/MIT_2006.

● TUDelft iGEM Team. (2008). TUDelft 2008. Retrieved on March 31th, 2014, from: https://2008.igem.org/Team:TUDelft.

● VictoriaBC. (2009). VictoriaBC 2009. Retrieved on March 31th, 2014, from: https://2009.igem.org/Team:VictoriaBC.

● Zubieta, Chole et al. (2003). Structural Basis for Substrate Recognition in the Salicylic Acid Carboxyl Methyltransferase Family. Manuscript submitted for publication. Retrieved from www.plantcell.org.


iGEM CIDEB 2014 - Footer

Retrieved from "http://2014hs.igem.org/Team:CIDEB-UANL_Mexico/project_aroma"