Difference between revisions of "Team:Rice/Wet Lab"

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         <div class = "h1" style="color:white">Introduction</div>
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         <div class = "h1" style="color:white">Overview</div>
 
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       <div class = "para">We facilitated a Building with Biology event at the Children’s Museum of Houston. Through hands-on activities like extracting DNA from wheat germ and helping kids to design their own superorganisms, we had the opportunity to introduce young children to the exciting field of synthetic biology and to clear up a few of their parents’ misconceptions.
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       <div class = "para"> Photoacoustic imaging is a technique in which contrast agents absorb photon energy and emit signals that can be analyzed by ultrasound. Currently, photoacoustics is used to image blood vessels because heme is a natural contrast agent found in blood. Photoacoustic imaging also provides a non-invasive alternative to current diagnostic tools used to detect internal tissue inflammation. In previous literature, hypoxia and nitric oxide have both been discovered to molecularly indicate gut inflammation, and iRFP670, 713, anacy and cyan have been found to emit wavelengths that are different from heme and can penetrate tissue with near-infrared wavelengths. Therefore, our goal is to report inflammation and cancer in the gut through photoacoustic imaging of engineered E. coli that express bacterial pigment violacein, as well as near-infrared fluorescent proteins iRFP670 and iRFP713.</div>
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         <div class = "h1" style="color:white">Background</div>
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         <div class = "h1" style="color:white">Arabinose Induced iRFP 670 and 713 Fluorescence</div>
 
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       <div class = "para" style="text-align:left;width: 565px;">The Building with Biology project  is dedicated to spreading STEM learning and discussion about the technological and societal implications of synthetic biology through public and scientist dialogue. Successful applicants receive a physical kit with all of the supplies necessary to host an event with six hands-on activities.
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pBAD is a very well-characterized expression system in E. coli. pBAD normally works by arabinose induction: araC, a constitutively produced transcription regulator, changes form in the presence of arabinose sugar, allowing for the activation of promoter pBAD. Therefore, we formed genetic circuits consisting of the pBAD expression system and iRFP670 and 713 to test the inducibility of our iRFPs. <br><br>
 
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        <li><div class = "para">Bio Bistro: Decide what current and future synthetic biology-based food products you would, would not, or might eat.</div></li>
 
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        <li><div class = "para">Kit of Parts: Solve challenges by building a model cell with standardized genetic parts (like BioBricks).</div></li>
 
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        <li><div class = "para">See DNA: Extract visible DNA from wheat germ and create necklaces to display your own sample of wheat germ DNA.</div></li>
 
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        <li><div class = "para">Super Organisms: Design a superhero to rescue a person falling from a tall building and then use that same creative engineering process to design a single-celled organism to clean up an oil spill.</div></li>
 
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        <li><div class = "para">Tech Tokens: Consider the potential advantages and disadvantages of various areas of synthetic biology research, before investing in them with "tech tokens."</div></li>
 
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        <li><div class = "para">VirEx Delivery: Explore the potential for engineered viruses to deliver beneficial, targeted genetic information to sites throughout the body.</div></li>
 
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         <div class = "h1" style="color:white">Nitric-oxide-induced Fluorescence</div>
 
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The next step was to test the nitric oxide induction of iRFP fluorescence. We used a genetic circuit consisting of a constitutive promoter that always expresses Part:BBa_K554003, which encodes for the expression of a SoxR. In the presence of nitric oxide, SoxR changes form to activate the promoter SoxS, which in turn is supposed to activate the expression of the iRFPs. Thus, for the next assay we added DETA/NO, a nitric oxide adduct in the presence of water.  
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  <div class = "para" id = "defaulti" style="display:box">Above is a timeline for Building with Biology, hover over different sections to learn more!</div>
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  <div class = "para" id = "onei" style="display:none">Volunteer orientation at the Children’s Museum & Building with Biology training</div>
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  <div class ="para" id = "twoi"style="display:none">Studying the background materials for our respective stations and practicing our presentations.</div>
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  <div class = "para" id = "sixi"style="display:none">Debrief</div>
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Revision as of 02:51, 22 November 2016















Overview


Photoacoustic imaging is a technique in which contrast agents absorb photon energy and emit signals that can be analyzed by ultrasound. Currently, photoacoustics is used to image blood vessels because heme is a natural contrast agent found in blood. Photoacoustic imaging also provides a non-invasive alternative to current diagnostic tools used to detect internal tissue inflammation. In previous literature, hypoxia and nitric oxide have both been discovered to molecularly indicate gut inflammation, and iRFP670, 713, anacy and cyan have been found to emit wavelengths that are different from heme and can penetrate tissue with near-infrared wavelengths. Therefore, our goal is to report inflammation and cancer in the gut through photoacoustic imaging of engineered E. coli that express bacterial pigment violacein, as well as near-infrared fluorescent proteins iRFP670 and iRFP713.


Arabinose Induced iRFP 670 and 713 Fluorescence


pBAD is a very well-characterized expression system in E. coli. pBAD normally works by arabinose induction: araC, a constitutively produced transcription regulator, changes form in the presence of arabinose sugar, allowing for the activation of promoter pBAD. Therefore, we formed genetic circuits consisting of the pBAD expression system and iRFP670 and 713 to test the inducibility of our iRFPs.



Nitric-oxide-induced Fluorescence


The next step was to test the nitric oxide induction of iRFP fluorescence. We used a genetic circuit consisting of a constitutive promoter that always expresses Part:BBa_K554003, which encodes for the expression of a SoxR. In the presence of nitric oxide, SoxR changes form to activate the promoter SoxS, which in turn is supposed to activate the expression of the iRFPs. Thus, for the next assay we added DETA/NO, a nitric oxide adduct in the presence of water.


Significance


The afternoon we spent at the Children’s Museum was particularly rewarding because we had the opportunity to interact with a cross section of the general public. The attendees had not come to the museum expecting to learn about synthetic biology, but they were eager to actively participate in the activities and to engage in conversations about the potential impact of the field. We were reminded by their vehement objections to buzzwords like “GMOs” of the significant resistance that synthetic biology still faces.