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− | {{NAIT_Edmonton/CSS2}} | + | {{NAIT_Edmonton/CSS6}} |
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| <link href='http://fonts.googleapis.com/css?family=Source+Sans+Pro' rel='stylesheet' type='text/css'> | | <link href='http://fonts.googleapis.com/css?family=Source+Sans+Pro' rel='stylesheet' type='text/css'> |
| + | <script type="text/javascript" src="https://ajax.googleapis.com/ajax/libs/jquery/1.6.2/jquery.min.js"></script> |
| + | <script type="text/javascript" src="https://2015.igem.org/Team:NAIT_Edmonton/jquery.flexslider-min?action=raw&ctype=text/javascript"></script> |
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− | }
| + | <div tabindex="0" class="onclick-menu"> |
| + | <ul class="onclick-menu-content" style="list-style:none;"> |
| + | <li><a href="https://2015.igem.org/Team:NAIT_Edmonton">home</a></li> |
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| + | <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Human">human practices</a></li> |
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− | </style> | + | <div class="navigation"> |
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| <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Team">team</a> | | <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Team">team</a> |
| <ul> | | <ul> |
| <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Bios">bios</a></li> | | <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Bios">bios</a></li> |
| <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Attributions">attributions</a></li> | | <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Attributions">attributions</a></li> |
− | <li><a href="https://igem.org/Team.cgi">official team profile</a> | + | <li><a href="https://igem.org/Team.cgi?id=1787">official team profile</a> |
| </ul> | | </ul> |
| </li> | | </li> |
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| <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Project">project</a> | | <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Project">project</a> |
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− | <div class="header_right">
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− | <ul>
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| <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Human">human practices</a> | | <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Human">human practices</a> |
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| <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Practices">policy and practices</a></li> | | <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Practices">policy and practices</a></li> |
| <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Collaborations">collaborations</a></li> | | <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Collaborations">collaborations</a></li> |
− | <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Entrepreneurship">entrepreneurship</a></li> | + | |
| </ul> | | </ul> |
| </li> | | </li> |
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| <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Notebook">notebook</a> | | <li><a href="https://2015.igem.org/Team:NAIT_Edmonton/Notebook">notebook</a> |
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| <style type="text/css"> | | <style type="text/css"> |
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| .top_slogan {text-align:center; font-family: 'Source Sans Pro', sans-serif; color:#0D4D8C; font-size:30px; padding: 40px 0px 40px 0px; font-style:strong; line-height:40px;} | | .top_slogan {text-align:center; font-family: 'Source Sans Pro', sans-serif; color:#0D4D8C; font-size:30px; padding: 40px 0px 40px 0px; font-style:strong; line-height:40px;} |
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| + | width: 580px; |
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| + | font-family: Arial, Helvetica sans-serif; |
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| box-sizing:border-box; | | box-sizing:border-box; |
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| text-align:center; | | text-align:center; |
| font-weight: 600; | | font-weight: 600; |
− | font-family:Arial, Helvetica, sans-serif; | + | font-family:'Source Sans Pro', sans-serif; |
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| .accordion-section-title.active, .accordion-section-title:hover { | | .accordion-section-title.active, .accordion-section-title:hover { |
| background:#9BD1EE; | | background:#9BD1EE; |
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| /* Type */ | | /* Type */ |
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| padding:20px 80px 20px 80px; | | padding:20px 80px 20px 80px; |
| display:none; | | display:none; |
− | font-size:15px; | + | font-size:18px; |
| } | | } |
| + | |
| + | .protocol {background-color:#6087c2; width:390px;margin:0 auto; -webkit-border-radius: 5px; |
| + | -moz-border-radius:5px; -ms-border-radius:5px; -0-border-radius:5px; |
| + | border-radius: 5px;padding:15px;} |
| + | .protocol:hover {background-color:#56aacc; width:390px;margin:0 auto; -webkit-border-radius: 5px; |
| + | -moz-border-radius:5px; -ms-border-radius:5px; -0-border-radius:5px; |
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| </style> | | </style> |
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− | <center><div class="top_slogan">The Project</div></center> | + | <center><div class="top_slogan">Development and Characterization of Protein Motifs to Generate Colours upon Interaction with Silver Staining Reagents</div></center> |
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− | <h2>Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS PAGE) is one of the most prominent techniques used in protein research. However, one of its limitations, more specifically the loss of an accurate point of reference post staining, is attributed to inefficiency and the wasting of time during the research process. From past observations, some proteins inherently produce certain colours post-silver staining. We hypothesized that certain configurations of amino acids produces specific colours when reacting with the reagents of silver staining. In the hopes of creating a new molecular weight ladder, our team designed sequences to code for novel proteins that produce colour. Coloured proteins can also pave the way for new types of colorimetric assays and a less expensive and more accessible counterpart to antibody tags in the future.</h2> <br> | + | <h2><b>Abstract: </b>SDS-PAGE is a very popular technique used to separate proteins based on their size. Embedded proteins, invisible to the naked eye, are then visualized by staining. Among the various staining techniques, silver staining is easy to perform and highly sensitive. However, the outcome is a series of monochromatic protein bands. Previously, we observed that some proteins inherently produce different hues post-staining. We hypothesized that specific amino acid configurations yield coloured bands after reacting with silver staining reagents. To test our hypothesis, we created numerous amino acid motifs to elucidate the sequences that would generate specific colours following silver staining. Our findings will let us generate a molecular weight marker with the innate capacity of providing users colour-coded bands post-staining without the use of impregnating dyes. Our technology will also pave the way for new types of colorimetric assays using synthetic proteins</h2> <br> |
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| <div class="accordion"> | | <div class="accordion"> |
| <div class="accordion-section"> | | <div class="accordion-section"> |
− | <a class="accordion-section-title" href="#accordion-1">Background</a> | + | <a class="accordion-section-title" href="#accordion-1" style="background-color:#f96040">Background</a> |
| <div id="accordion-1" class="accordion-section-content"> | | <div id="accordion-1" class="accordion-section-content"> |
| The structural and functional study of the proteins expressed by a genome is | | The structural and functional study of the proteins expressed by a genome is |
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| <br> | | <br> |
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− | <center><img src="http://upload.wikimedia.org/wikipedia/commons/4/46/SDS-PAGE_Electrophoresis.png" width="750px"></center> | + | <center><img src="https://static.igem.org/mediawiki/2015/b/bb/NAIT_SDSPAGE.png" width="750px"></center> |
| + | <p style="font-size:10px">Source: http://upload.wikimedia.org/wikipedia/commons/4/46/SDS-PAGE_Electrophoresis.png</p> |
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| <br> | | <br> |
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− | <center><img src="http://www.bio-rad.com/webroot/web/images/lsr/products/electrophoresis/product_overlay_content/global/lsr_biosafe_coomasie_gel.jpg"></center> | + | <center><img src="https://static.igem.org/mediawiki/2015/b/b0/Lsr_biosafe_coomasie_gel.jpg"></center> |
| + | <p><font size="1px">[1] Bio-rad.com, 'Coomassie Stains', 2015. [Online]. Available: http://www.bio-rad.com/webroot/web/images/lsr/products/electrophoresis/product_overlay_content/global/lsr_biosafe_coomasie_gel.jpg. [Accessed: 14- Jun- 2015].</font></p> |
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| <div class="accordion-section"> | | <div class="accordion-section"> |
− | <a class="accordion-section-title" href="#accordion-2">The Problem</a> | + | <a class="accordion-section-title" href="#accordion-2" style="background-color:#FBB252">The Problem</a> |
| <div id="accordion-2" class="accordion-section-content"> | | <div id="accordion-2" class="accordion-section-content"> |
− | <p>Difficulties with silver staining arise when the molecular weight markers are re- | + | <p>However, there are limitations to using the silver staining technique. The primary issue with the technique is that the users lose the ability to use the color coded molecular weight marker as a reference post-staining. To combat this issue, researchers poke holes into the polyacrylamide gel so that they can retain their molecular weight ladder reference points. However, by doing this we ruin the integrity of the gel making the staining process much more likely to damage or destroy the fragile gel. </p> |
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− | colored golden-brown in the staining process. Markers offer evenly distributed proteins
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− | that show bands of equal intensity and known size. Researchers can compare these
| + | <center><img src="https://static.igem.org/mediawiki/2015/a/a9/NAIT_SilverStain.jpeg" width="750px"> |
| + | <p style="font-size:10px">Source:http://labs.mmg.pitt.edu/gjoerup/silver_stain.jpg</p> |
| + | </center><br> |
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− | bands with their sample and identify the protein they are looking for based on its size. A
| + | <p>Researchers who have used the technique in the past have also expressed concerns on the time it takes to develop the gel and stain; the possibility of “over-staining” the gel, and the generation of false positives due to the sensitivity of the technique. (An example would be the solutions reacting with particles in the air to provide an unneeded background on the gel) </p> |
− | | + | |
− | subset of these markers has color-coded standard proteins to facilitate the identification
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− | | + | |
− | of each band. Post-silver staining, the users lose the ability to use the color code as a | + | |
− | | + | |
− | reference.</p>
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− | | + | |
− | <br>
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− | | + | |
− | <center><img src="http://labs.mmg.pitt.edu/gjoerup/silver_stain.jpg" width="750px"></center>
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| </div><!--end .accordion-section-content--> | | </div><!--end .accordion-section-content--> |
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| <div class="accordion-section"> | | <div class="accordion-section"> |
− | <a class="accordion-section-title" href="#accordion-3">Our Goal</a> | + | <a class="accordion-section-title" href="#accordion-3" style="background-color: #f7e133">Our Goal and Solution</a> |
| <div id="accordion-3" class="accordion-section-content"> | | <div id="accordion-3" class="accordion-section-content"> |
− | <p>Our goal is to develop a marker that, when interacting with the reagents used in | + | <p>Our goal is to develop a marker that, when interacting with the reagents used in the staining protocol, will stain in colour and in specific positions. This technology will aid in the identification of the protein(s) of interest post-staining. In order to do so, we will investigate how specific amino acids react with silver staining reagents. After determining specific motifs, we aim to create novel proteins that contain an excess of that motif, particular amino acid and/or chemical modifications that will generate a specific color after treating it with silver staining reagents. To obtain such proteins, we will introduce novel nucleotide sequences into a plasmid by in vitro transcription translation. <i>E. coli</i> cells with expression vectors will then be transformed with the new plasmids.</p> <br> |
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− | the staining protocol, will develop colour bands in specific positions so as to help in the | + | <p>By testing different techniques to develop and stain the gels we hope to see if the expressed concerns are still an issue, namely time taken to develop the gels and stain them effectively. To investigate this we will use BioRad’s recommended protocol, our protocol developed in-house, and a technique documented in <i>Electrophoresis.</i></p> |
− | | + | |
− | identification of the protein(s) of interest post-staining. In order to do so, investigation of
| + | |
− | | + | |
− | how specific amino acids react with silver staining reagents is underway by our team.
| + | |
− | | + | |
− | This will have as an outcome the creation of novel proteins that contain an excess of a
| + | |
− | | + | |
− | particular amino acid and/or chemical modifications that will generate a specific colour
| + | |
− | | + | |
− | after treating it with silver staining reagents. To obtain such proteins, the introduction of
| + | |
− | | + | |
− | novel nucleotide sequences into a plasmid would be done first by in vitro transcription
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− | | + | |
− | translation and later by transforming E. coli cells with expression vectors.</p>
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| </div><!--end .accordion-section-content--> | | </div><!--end .accordion-section-content--> |
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| <div class="accordion-section"> | | <div class="accordion-section"> |
− | <a class="accordion-section-title" href="#accordion-4">Our Solution</a> | + | <a class="accordion-section-title" href="#accordion-5" style="background-color:#afcf64">Why Does this Matter?</a> |
− | <div id="accordion-4" class="accordion-section-content">
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− | <p>Design novel protein sequences that will stain in colour.</p>
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− | | + | |
− | </div><!--end .accordion-section-content-->
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− | </div><!--end .accordion-section-->
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− | <a class="accordion-section-title" href="#accordion-5">Why Does this Matter?</a>
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| <div id="accordion-5" class="accordion-section-content"> | | <div id="accordion-5" class="accordion-section-content"> |
| <p>With the current technique used today, researchers poke holes into the PA gel so that they can retain their molecular weight ladder reference points. Not only does this take time to do, but it also ruins the integrity of the gel making the staining process much more likely to damage the fragile gel.</p> | | <p>With the current technique used today, researchers poke holes into the PA gel so that they can retain their molecular weight ladder reference points. Not only does this take time to do, but it also ruins the integrity of the gel making the staining process much more likely to damage the fragile gel.</p> |
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− | .footer {position: relative; padding:20px 0 40px 0; margin: -25px 0 0 0; clear:both; width:100%; background-color:#0D4D8C; height:25px;}
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− | .footer_content {margin:auto;width:100%;}
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− | .footer_left { float:left; padding:0 0 0 10%;}
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− | .footer_right{ float:right; padding:0 10% 0 0;}
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− | ul.social_icons { margin:0px; padding:0px; list-style:none;}
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− | ul.social_icons li{ float:left;}
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− | ul.social_icons li a{width:30px;height:30px;display:block;border-radius:15px;-moz-border-radius:15px;-webkit-border-radius:15px;-khtml-border-radius:15px;float:left;margin:0px 0 0px 5px;background-color:#ADD3F0;text-align:center;}
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− | ul.social_icons li a#top{width:30px;height:30px;display:block;border-radius:15px;-moz-border-radius:15px;-webkit-border-radius:15px;-khtml-border-radius:15px;float:left;margin:0px 0 0px 5px;background-color:#ADD3F0;text-align:center;}
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− | ul.social_icons li a:hover{ background-color:#5AB3E3;}
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− | ul.social_icons li a img{ width:60%; display:block; margin:5px auto;}
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− |
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− | ul.footer_menu{ padding:10px 0 0 0; margin:0px; list-style:none;}
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− | ul.footer_menu li{ float:left; padding:0 15px 0 0;}
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− | ul.footer_menu li a{ background:url(https://static.igem.org/mediawiki/2015/1/17/NAIT_Bullet.png) no-repeat left; padding:0 0 0 12px;}
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− |
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− |
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− | </style>
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| | | |
| </body> | | </body> |
− |
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− | <!----------
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− |
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− | _ __ _
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− | ;' '',)
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− | /;6 , ;/
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− | (Y)_:., |
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− | `-', :; \
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− | |; ,.:\
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− | /:.; ;;)
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− | |:;,.'| :/
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− | / |: / ; /
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− | /:;\ `| "//
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− | /_,: | |./,|
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− | _/: \.'|,|/| |
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− | /:.,:.|,|"| |:|
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− | eduardo |',:| \_ \ |_|;\_
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− | /;\_ /\_)) \_))\_))
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− | (;(________
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− | '''''`'''~`
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− |
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− | ------------------>
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− |
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| | | |
| </html> | | </html> |