Difference between revisions of "Team:IIT Delhi/Model"

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     </div>
 
     </div>
 
   </div>
 
   </div>
 
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<div class="dropdownsakks">
 
         <div class= "borderYtoX">
 
         <div class= "borderYtoX">
   <a href="https://2018.igem.org/Team:IIT_Delhi/Results" >Results</a>
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   <a href="javascript:void(0)" >Results</a>
 
     </div>
 
     </div>
  
          
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  <div class="dropdownsakks-content"  style="margin-top: 66px;">
 +
      <a href="https://2018.igem.org/Team:IIT_Delhi/Demonstrate">Demonstrate</a>
 +
      <a href="https://2018.igem.org/Team:IIT_Delhi/InterLab">Interlab</a>
 +
     
 +
    </div>
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     </div>
 
     </div>
 
      
 
      
        <a href="https://2018.igem.org/Team:IIT_Delhi/Results" style="padding-bottom: 15px;padding-top: 15px;" >Results</a>
 
       
 
 
     <button class="overlayved-btn" onclick="myFunction2()" style="padding-bottom: 15px;">Parts
 
     <button class="overlayved-btn" onclick="myFunction2()" style="padding-bottom: 15px;">Parts
 
       <i class="fa fa-caret-right"></i>   
 
       <i class="fa fa-caret-right"></i>   
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     <a href="https://2018.igem.org/Team:IIT_Delhi/Basic_Part" style="font-size: 15px;font-size: 15px; padding-bottom: 5px; padding: 10px; padding-top: 3px;" >Basic Parts</a>
 
     <a href="https://2018.igem.org/Team:IIT_Delhi/Basic_Part" style="font-size: 15px;font-size: 15px; padding-bottom: 5px; padding: 10px; padding-top: 3px;" >Basic Parts</a>
 
     <a href="https://2018.igem.org/Team:IIT_Delhi/Composite_Part" style="font-size: 15px;font-size: 15px; padding: 10px; padding-bottom: 5px; padding-top: 5px;" >Composite Parts</a>
 
     <a href="https://2018.igem.org/Team:IIT_Delhi/Composite_Part" style="font-size: 15px;font-size: 15px; padding: 10px; padding-bottom: 5px; padding-top: 5px;" >Composite Parts</a>
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    </div>
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 +
       
 +
    <button class="overlayved-btn" onclick="myFunction6()" style="padding-bottom: 15px;">Results
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      <i class="fa fa-caret-right"></i> 
 +
    </button>
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 +
    <div class="dropdownsakks-container" id="mydropdownsakks6" style= "font-style: italic;">
 +
    <a href="https://2018.igem.org/Team:IIT_Delhi/Demonstrate" style="font-size: 15px;font-size: 15px; padding-bottom: 5px; padding: 10px; padding-top: 3px;" >Demonstrate</a>
 +
    <a href="https://2018.igem.org/Team:IIT_Delhi/InterLab" style="font-size: 15px;font-size: 15px; padding: 10px; padding-bottom: 5px; padding-top: 5px;" >Interlab</a>
 
            
 
            
 
     </div>
 
     </div>
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</section>
 
</section>
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<div class="bgimg-1">
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  <div class="caption">
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    <span class="border" >Text anyone?</span>
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  </div>
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</div>
  
 
 
  
 
<div style="color: #777;background-color:white;text-align:center;padding:50px 80px;text-align: justify;">
 
<div style="color: #777;background-color:white;text-align:center;padding:50px 80px;text-align: justify;">
  <!--<h3 style="text-align:center;text-decoration:hotpink double underline">Safety Form</h3>-->
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   <p style="font-size: 27px; font-family: 'Roboto', sans-serif;font-weight:550;">We at iGEM understand the necessity of working in a safe environment. A safe environment ensures a non-hesitant mind that designs more efficient biological devices.
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   <p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814001 = pLTL
 
     <br>
 
     <br>
 
       </p>
 
       </p>
       <p style="font-size:20px;">About lab safety and training:</p>
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       <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;"> The pLTL (Lac-Tet-Lac) is a hybrid promoter. It is a promoter composed of the operator sequences of pTet and the pLac promoter. There are multiple benefits of using the pLTL promoter.
 +
</p>
 
      
 
      
 
       <ul style="font-size:20px;" >
 
       <ul style="font-size:20px;" >
       <li style="font-family: 'Lato', sans-serif;font-weight:400;">Lab access and rules (including appropriate clothing, eating and drinking, etc.).</li>
+
       <li style="font-family: 'Lato', sans-serif;font-weight:400;">The hybrid pLTL shows almost complete repression on being repressed, and on induction (by IPTG and/or aTc), the hybrid pLTL promoter shows a 1.4-2 times higher expression.</li>
      <li style="font-family: 'Lato', sans-serif;font-weight:400;">Responsible individuals (such as lab or departmental specialist or institutional biosafety officer).</li>
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       <li style="font-family: 'Lato', sans-serif;font-weight:400;">The hybrid pLTL promoter also permits flexible gene expression because it can be utilized under either or both repression controls (LacI and TetR) simultaneously.</li>
      <li style="font-family: 'Lato', sans-serif;font-weight:400;">Differences between biosafety levels.</li>
+
   
       <li style="font-family: 'Lato', sans-serif;font-weight:400;">Biosafety equipment (such as biosafety cabinets).</li>
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    <li style="font-family: 'Lato', sans-serif;font-weight:400;">Good microbial technique (such as lab practices).</li>
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    <li style="font-family: 'Lato', sans-serif;font-weight:400;">Disinfection and sterilization.</li>
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      <li style="font-family: 'Lato', sans-serif;font-weight:400;">Emergency procedures.</li>
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          <li style="font-family: 'Lato', sans-serif;font-weight:400;">Chemicals, fire and electrical safety.</li>
+
 
     </ul>
 
     </ul>
 +
<br>
  
      
+
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814008 = rrnB T1 Terminator + T7Te Terminator
 +
     <br>
 +
      </p>
 +
      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;"> Our part is double terminator composed of rrnB T1 Terminator(BBa B0010) and T7 Te Terminator(BBa B0012). Both of these are forward terminators that are extensively used in E. coli.
 +
</p>
 +
<br>
 +
 
 +
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814009 = mKate2
 +
 
 +
    <br>
 +
      </p>
 +
      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;"> Mkate2 - mKate2 is a red fluorescent protein derived from Entacmaea quadricolor.  It possesses fluorescence with excitation maxima at 588 nm and emission maxima at 588 and 633 nm, mKate2 is almost 3-fold brighter than mKate. mKate2 can be used in  labelling applications along with blue, cyan, green, yellow, and red fluorescent dyes. Its high pH-stability with pKa=5.4 makes it useful for imaging in acidic organelles, such as late and recycling endosomes and lysosomes.
 +
 
 +
</p>
 +
<br>
 +
 
 +
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814010 = rrnB T1 Terminator
 +
 
 +
    <br>
 +
      </p>
 +
    <br>
 
</div>
 
</div>
  
<div class="bgimg-1">
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<div class="bgimg-2">
 
   <div class="caption">
 
   <div class="caption">
     <span class="border" >Bio-safety Rules and Regulations</span>
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     <span class="border" >Text anyone?</span>
 
   </div>
 
   </div>
 
</div>
 
</div>
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<div style="position:relative;">
 
<div style="position:relative;">
 
   <div style="color: #777;background-color:white;text-align:center;padding:50px 80px;text-align: justify;">
 
   <div style="color: #777;background-color:white;text-align:center;padding:50px 80px;text-align: justify;">
  <ul style="font-size:20px;">
 
<li style="font-family: 'Lato', sans-serif;font-weight:400;"> iGEM IIT Delhi has always been following all biosafety rules of our institution and concerned biosafety laws of our country. </li>
 
<li style="font-family: 'Lato', sans-serif;font-weight:400;">EHLSU (Environmental Health and Lab Safety Unit) is responsible for holding up all the safety guidelines in all the labs (including ours) at the institution. The guidelines can be found on the link -<br>
 
        <a href="http://www.dbtindia.nic.in/wp-content/uploads/Draft-Biosafety-Regulations-and-Biocontainment-Guidelines-2017-FF.pdf">http://www.dbtindia.nic.in/wp-content/uploads/Draft-Biosafety-Regulations-and-Biocontainment-Guidelines-2017-FF.pdf</a> </li>
 
<li style="font-family: 'Lato', sans-serif;font-weight:400;">The safety guidelines for all the labs in our region (North India) are declared and monitored by EHLSU, whose rules can be found on the following link - <br>
 
        <a href="http://www.dbtindia.nic.in/wp-content/uploads/Draft-Biosafety-Regulations-and-Biocontainment-Guidelines-2017-FF.pdf">http://www.dbtindia.nic.in/wp-content/uploads/Draft-Biosafety-Regulations-and-Biocontainment-Guidelines-2017-FF.pdf</a> </li>
 
<li style="font-family: 'Lato', sans-serif;font-weight:400;">iGEM IIT Delhi did not conduct any experiment with human subjects (including non-invasive experiments, such as surveys), for the surveys conducted we have followed the rules of our institution. </li>
 
  </ul>
 
 
 
    
 
    
 +
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814007 = sfGFP_ssrA
 +
 +
 +
    <br>
 +
      </p>
 +
      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">Our part consists of sfGFP appended with a ssrA(LVA) deg-tag. It has been codon optimised for E. coli.  This part is useful as it has high intensity as compared to other gfp variants as well as faster degradation rates and shorter reporter lifetimes.
 +
 +
 +
</p>
 +
 +
<p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">Superfolder GFP is a basic (constitutively fluorescent) green fluorescent protein derived from Aequorea victoria. It has an emission wavelength of 510 nm and excitation wavelength of 485nm. It has a robust folding characteristic. The superfolder mutations also make the folding of GFP tolerant of mutations that would otherwise reduce the folding yield of GFP.
 +
 +
 +
</p>
 +
 +
<p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">Our ssrA degtag is the gfp(LVA) that has a single (A → G) point mutation in nucleotide 349, resulting in an Asp117 → Gly117 (D117G) amino acid change. This point mutation does not appear to change the fluorescence spectrum of gfp(LVA). The LVA tag has been reported to lead to fast protein degradation, degrading GFP with rate -0.018 per minute. This corresponds to in vivo half-lives of mature Gfp(LVA) of approximately 40 min.
 +
 +
 +
 +
</p>
 +
 +
<p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">References</p>
 +
<ol style="font-size: 20px;">
 +
 +
<li style="font-family: 'Lato', sans-serif;font-weight:400;">Pédelacq JD, Cabantous S, Tran T, Terwilliger TC, Waldo GS. Engineering and characterization of a superfolder green fluorescent protein. Nature biotechnology. 2006 Jan;24(1):79.</li>
 +
 +
<li style="font-family: 'Lato', sans-serif;font-weight:400;">Andersen, J. B. et al. New unstable variants of green fluorescent protein for studies of transient gene expression in bacteria. Appl. Environ. Microbiol. 64, 2240–6 (1998).
 +
</li>
 +
 +
 +
  </ol>
 +
 +
<br>
 +
 +
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814011 = attP TP901-1
 +
 +
 +
    <br>
 +
      </p>
 +
      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">AttP TP901-1 - It is the AttP site of TP901-1 integrase (serine based recombinase enzyme). TP901-1 integrase is an enzyme that has been isolated from TP901-1 phage. It is responsible for catalysing site-specific recombination at AttP and AttB sites. The recombination mechanism depends on the orientation of the AttP and AttB sites. These enzymes help the virus integrate its DNA into the bacterial genome. TP901-1 is widely used in the construction of logic gates and modification of DNA sequences.
 +
 +
 +
</p>
 +
 
 +
<br>
 +
 +
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814012 = BxbI Integrase + ssrA(LVA) deg tag
 +
 +
 +
    <br>
 +
      </p>
 +
      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">BxbI Integrase triggers attP × attB recombination. The product of attP × attB recombination is an integrated prophage flanked by two new recombination sites, attL and attR, each containing half sites derived from attP and attB. In the absence of accessory factors the integrases mediate unidirectional recombination between attP and attB with greater than 80% efficiency. In the presence of a phage-encoded accessory protein, the recombination directionality factor (RDF) the attP × attB recombination is inhibited and the attL × attR recombination is stimulated.
 +
Bxb1 integrase yields approximately two-fold more recombinants  and displays about two fold less damage to the recombination sites than other phage-encoded serine integrases.
 +
 +
Our BxbI Integrase has a ssrA deg tag attached to it for faster degradation rates.
 +
 +
 +
 +
</p>
 +
 +
<br>
 +
 +
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814013 = attB TP901-1
 +
 +
 +
    <br>
 +
      </p>
 +
      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">AttB TP901-1 -  It is the AttB site of TP901-1 integrase (serine based recombinase enzyme). TP901-1 integrase is an enzyme that has been isolated from TP901-1 phage. It is responsible for catalysing site-specific recombination at AttP and AttB sites. The recombination mechanism depends on the orientation of the AttP and AttB sites. These enzymes help the virus integrate its DNA into the bacterial genome. TP901-1 is widely used in the construction of logic gates and changing DNA sequence.
 +
 +
 +
 +
</p>
 +
 +
<br>
 +
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814014 = complement of B0034, RBS on the antisense strand, twin exists
 +
    <br>
 +
      </p>
 +
<br>
 +
 +
 
   </div>
 
   </div>
 
</div>
 
</div>
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<div class="bgimg-3">
 
<div class="bgimg-3">
 
   <div class="caption">
 
   <div class="caption">
     <span class="border">General Safety Rules of the lab</span>
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     <span class="border">Some random text </span>
 
   </div>
 
   </div>
 
</div>
 
</div>
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<div style="position:relative;">
 
<div style="position:relative;">
 
   <div style="color: #777;background-color:white;text-align:center;padding:50px 80px;text-align: justify;">
 
   <div style="color: #777;background-color:white;text-align:center;padding:50px 80px;text-align: justify;">
     <ul style="font-size:20px;">   
+
      
        <li style="font-family: 'Lato', sans-serif;font-weight:400;">Instructions are strictly followed</li>
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<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814015 = pLac(lambda) hybrid
        <li style="font-family: 'Lato', sans-serif;font-weight:400;">Personal protection equipment is must and hair should be tied</li>
+
 
        <li style="font-family: 'Lato', sans-serif;font-weight:400;">Working areas and common work areas are kept tidy and regularly wiped with cleansing agents like ethanol</li>
+
 
        <li style="font-family: 'Lato', sans-serif;font-weight:400;">Pipette tips, media, eppendorf tubes are autoclaved and Glassware; tools are cleaned with ethanol before usage</li>
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    <br>
        <li style="font-family: 'Lato', sans-serif;font-weight:400;">Handwash is a must after entering and before leaving the lab.</li>
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      </p>
        <li style="font-family: 'Lato', sans-serif;font-weight:400;">Gloves are used while handling carcinogenic agents like EtBr and high temperature objects. </li>
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      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">pLac - Lambda hybrid - is a hybrid promoter consisting of Lac and Lambda operator sites in the core region. This hybrid promoter can be induced in the presence of  IPTG or
        <li style="font-family: 'Lato', sans-serif;font-weight:400;">There is a separate work bench for experiments in which toxic chemicals (e.g. Electrophoresis, Etbr).</li>  
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</p>
        <li style="font-family: 'Lato', sans-serif;font-weight:400;">Eating or drinking is prohibited in the lab.</li>
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<br>
        <li style="font-family: 'Lato', sans-serif;font-weight:400;">Spills are immediately cleaned.</li>
+
 
        <li style="font-family: 'Lato', sans-serif;font-weight:400;">Lab is equipped with safety equipment and everyone is aware of its location (E.g. Fire extinguishers and first aid supplies).</li>
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<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814017 = attP Bxb1
        <li style="font-family: 'Lato', sans-serif;font-weight:400;">Appropriate protection is used while working with UV light(eye protection, gloves, lab coat).</li>
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        <li style="font-family: 'Lato', sans-serif;font-weight:400;">Lab waste is properly disposed (E.g. Biological cultures are disposed after addition of ethanol to kill microorganisms; Solids are kept out of the sink; Agarose gel, gloves, the hazardous wastes are disposed separately in the biohazard coloured bin).</li>
+
 
      </ul>
+
 
 +
    <br>
 +
      </p>
 +
      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">AttP Bxb1 - It is the AttP site of Bxb1 integrase (Serine based recombinase). Bxb1 integrase carries out site-specific recombination by catalysing unidirectional recombination to produce AttL and AttR sites from AttP and AttB. This switching capacity allows them to be used in the design of toggle switches.
 +
 
 +
 
 +
</p>
 +
<br>
 +
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814018 = attB Bxb1
 +
 
 +
 
 +
    <br>
 +
      </p>
 +
      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">AttB Bxb1 - It is the AttB site of Bxb1 integrase (Serine based recombinase). Bxb1 integrase carries out site-specific recombination by catalysing unidirectional recombination to produce AttL and AttR sites from AttP and AttB. This switching capacity allows them to be used in the design of logic gates.
 +
 
 +
 
 +
</p>
 +
<br>
 +
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814019 = P7 Promoter
 +
 
 +
 
 +
 
 +
 
 +
    <br>
 +
      </p>
 +
      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">Constitutive P7 promoter - It is the complement of the constitutive P7 promoter so as to initiate transcription in the reverse direction.
 +
 
 +
</p>
 +
 
 +
<br>
 +
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814021 = BxbI-Xis  + ssrA(LVA) deg tag------------
 +
 
 +
 
 +
    <br>
 +
      </p>
 +
      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">Xis-BxbI_ssrA deg tag - Consists of bxb1 excisionase followed by ssrA degradation tag. Bxb1-Xis  catalyses the conversion of AttL and AttR sites to AttP and AttB sites when expressed with Bxb1 integrase, thereby reverting the recombination caused by integrase. ssrA deg tag degrades the Bxb1-Xis formed as an increase in the amount of excisionase renders the system inefficient. This property allows Bxb1 to be used in the construction of logic gates.
 +
</p>
 +
 
 +
<br>
 +
 
 +
<p style="font-size: 30px; font-family: 'Roboto', sans-serif;font-weight:700;">BBa_K2814022 = attB BxbI (reverse orientation)
 +
 
 +
 
 +
    <br>
 +
      </p>
 +
      <p style="font-size:20px;font-family: 'Lato', sans-serif;font-weight:400;">AttB-BxbI (Reverse Orientation) - Contains AttB site of Bxb1 integrase in the reverse orientation (Reverse of  BBa_K2814018). Bxb1 integrase carries out site-specific recombination by catalysing unidirectional recombination to produce AttL and AttR sites from AttP and AttB. This switching capacity allows them to be used in the design of logic gates.  
 +
 
 +
</p>
 +
 
 +
<br>
 +
 
 +
 
 
   </div>
 
   </div>
 
</div>
 
</div>
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<div style="background-color: white">
 
<div style="background-color: white">
         <div  style="background-color: #555;font-size: 35px;text-align: center;font-family: sans-serif;"><h4 style="color: white">Contact us</h4></div>
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         <div  style="background-color: #555;color:black;font-size: 35px;text-align: center;font-family: sans-serif;"><h4 style="color: white">Contact us</h4></div>
             <div style="text-align: center;margin-bottom: -136px;"><h4 style="font-size: 50px;"><u class="fx" style="cursor: pointer;text-align: center">Address</u></h4><p style="text-align: center;margin-bottom: 16px">Undergraduate Laboratory<br>
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             <div style="text-align: center;margin-bottom: -136px;"><h4 style="color:black;font-size: 50px;"><u class="fx" style="cursor: pointer;text-align: center">Address</u></h4><p style="text-align: center;margin-bottom: 16px">Undergraduate Laboratory<br>
 
Department of Biotechnology and Biochemical Engineering, IIT Delhi</p>  
 
Department of Biotechnology and Biochemical Engineering, IIT Delhi</p>  
 
             </div>
 
             </div>
         <div style="margin-left: 75%;height: auto;max-height: inherit;width: auto;max-width: inherit;margin-bottom: 8px;"><i class="fa fa-facebook-official fx" style="font-size:90px;transition: 0.5s;transition-duration: 0.3s;transition-property: transform;margin-bottom: 30px;"></i></div>
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         <div style="margin-left: 75%;height: auto;max-height: inherit;width: auto;max-width: inherit;margin-bottom: 8px;"><a href="https://www.facebook.com/igemiitdelhi/" style="color:black;"><i class="fa fa-facebook-official fx" style="font-size:90px;transition: 0.5s;transition-duration: 0.3s;transition-property: transform;margin-bottom: 30px;"></i></a></div>
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Revision as of 01:01, 17 October 2018

iGEM IIT Delhi

Text anyone?

BBa_K2814001 = pLTL

The pLTL (Lac-Tet-Lac) is a hybrid promoter. It is a promoter composed of the operator sequences of pTet and the pLac promoter. There are multiple benefits of using the pLTL promoter.

  • The hybrid pLTL shows almost complete repression on being repressed, and on induction (by IPTG and/or aTc), the hybrid pLTL promoter shows a 1.4-2 times higher expression.
  • The hybrid pLTL promoter also permits flexible gene expression because it can be utilized under either or both repression controls (LacI and TetR) simultaneously.

BBa_K2814008 = rrnB T1 Terminator + T7Te Terminator

Our part is double terminator composed of rrnB T1 Terminator(BBa B0010) and T7 Te Terminator(BBa B0012). Both of these are forward terminators that are extensively used in E. coli.


BBa_K2814009 = mKate2

Mkate2 - mKate2 is a red fluorescent protein derived from Entacmaea quadricolor. It possesses fluorescence with excitation maxima at 588 nm and emission maxima at 588 and 633 nm, mKate2 is almost 3-fold brighter than mKate. mKate2 can be used in labelling applications along with blue, cyan, green, yellow, and red fluorescent dyes. Its high pH-stability with pKa=5.4 makes it useful for imaging in acidic organelles, such as late and recycling endosomes and lysosomes.


BBa_K2814010 = rrnB T1 Terminator


Text anyone?

BBa_K2814007 = sfGFP_ssrA

Our part consists of sfGFP appended with a ssrA(LVA) deg-tag. It has been codon optimised for E. coli. This part is useful as it has high intensity as compared to other gfp variants as well as faster degradation rates and shorter reporter lifetimes.

Superfolder GFP is a basic (constitutively fluorescent) green fluorescent protein derived from Aequorea victoria. It has an emission wavelength of 510 nm and excitation wavelength of 485nm. It has a robust folding characteristic. The superfolder mutations also make the folding of GFP tolerant of mutations that would otherwise reduce the folding yield of GFP.

Our ssrA degtag is the gfp(LVA) that has a single (A → G) point mutation in nucleotide 349, resulting in an Asp117 → Gly117 (D117G) amino acid change. This point mutation does not appear to change the fluorescence spectrum of gfp(LVA). The LVA tag has been reported to lead to fast protein degradation, degrading GFP with rate -0.018 per minute. This corresponds to in vivo half-lives of mature Gfp(LVA) of approximately 40 min.

References

  1. Pédelacq JD, Cabantous S, Tran T, Terwilliger TC, Waldo GS. Engineering and characterization of a superfolder green fluorescent protein. Nature biotechnology. 2006 Jan;24(1):79.
  2. Andersen, J. B. et al. New unstable variants of green fluorescent protein for studies of transient gene expression in bacteria. Appl. Environ. Microbiol. 64, 2240–6 (1998).

BBa_K2814011 = attP TP901-1

AttP TP901-1 - It is the AttP site of TP901-1 integrase (serine based recombinase enzyme). TP901-1 integrase is an enzyme that has been isolated from TP901-1 phage. It is responsible for catalysing site-specific recombination at AttP and AttB sites. The recombination mechanism depends on the orientation of the AttP and AttB sites. These enzymes help the virus integrate its DNA into the bacterial genome. TP901-1 is widely used in the construction of logic gates and modification of DNA sequences.


BBa_K2814012 = BxbI Integrase + ssrA(LVA) deg tag

BxbI Integrase triggers attP × attB recombination. The product of attP × attB recombination is an integrated prophage flanked by two new recombination sites, attL and attR, each containing half sites derived from attP and attB. In the absence of accessory factors the integrases mediate unidirectional recombination between attP and attB with greater than 80% efficiency. In the presence of a phage-encoded accessory protein, the recombination directionality factor (RDF) the attP × attB recombination is inhibited and the attL × attR recombination is stimulated. Bxb1 integrase yields approximately two-fold more recombinants and displays about two fold less damage to the recombination sites than other phage-encoded serine integrases. Our BxbI Integrase has a ssrA deg tag attached to it for faster degradation rates.


BBa_K2814013 = attB TP901-1

AttB TP901-1 - It is the AttB site of TP901-1 integrase (serine based recombinase enzyme). TP901-1 integrase is an enzyme that has been isolated from TP901-1 phage. It is responsible for catalysing site-specific recombination at AttP and AttB sites. The recombination mechanism depends on the orientation of the AttP and AttB sites. These enzymes help the virus integrate its DNA into the bacterial genome. TP901-1 is widely used in the construction of logic gates and changing DNA sequence.


BBa_K2814014 = complement of B0034, RBS on the antisense strand, twin exists


Some random text

BBa_K2814015 = pLac(lambda) hybrid

pLac - Lambda hybrid - is a hybrid promoter consisting of Lac and Lambda operator sites in the core region. This hybrid promoter can be induced in the presence of IPTG or


BBa_K2814017 = attP Bxb1

AttP Bxb1 - It is the AttP site of Bxb1 integrase (Serine based recombinase). Bxb1 integrase carries out site-specific recombination by catalysing unidirectional recombination to produce AttL and AttR sites from AttP and AttB. This switching capacity allows them to be used in the design of toggle switches.


BBa_K2814018 = attB Bxb1

AttB Bxb1 - It is the AttB site of Bxb1 integrase (Serine based recombinase). Bxb1 integrase carries out site-specific recombination by catalysing unidirectional recombination to produce AttL and AttR sites from AttP and AttB. This switching capacity allows them to be used in the design of logic gates.


BBa_K2814019 = P7 Promoter

Constitutive P7 promoter - It is the complement of the constitutive P7 promoter so as to initiate transcription in the reverse direction.


BBa_K2814021 = BxbI-Xis + ssrA(LVA) deg tag------------

Xis-BxbI_ssrA deg tag - Consists of bxb1 excisionase followed by ssrA degradation tag. Bxb1-Xis catalyses the conversion of AttL and AttR sites to AttP and AttB sites when expressed with Bxb1 integrase, thereby reverting the recombination caused by integrase. ssrA deg tag degrades the Bxb1-Xis formed as an increase in the amount of excisionase renders the system inefficient. This property allows Bxb1 to be used in the construction of logic gates.


BBa_K2814022 = attB BxbI (reverse orientation)

AttB-BxbI (Reverse Orientation) - Contains AttB site of Bxb1 integrase in the reverse orientation (Reverse of BBa_K2814018). Bxb1 integrase carries out site-specific recombination by catalysing unidirectional recombination to produce AttL and AttR sites from AttP and AttB. This switching capacity allows them to be used in the design of logic gates.


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