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

 
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#banner {
 
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background: url("https://static.igem.org/mediawiki/2017/5/5c/T--IIT_DELHI--modelling_banner.jpg") ;
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</head>
 
</head>
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<body class="landing .igem_iitd_delhi_body" background-color: black;>
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<!-- Header -->
 
<!-- Header -->
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     </button>
 
     </button>
 
     <div class="dropdown-content">
 
     <div class="dropdown-content">
−
       <a href="/Team:IIT_Delhi/Description">Description</a>
+
       <a href="/Team:IIT_Delhi/Description">Overview</a>
−
       <a href="/Team:IIT_Delhi/Results">Results</a>
+
       <a href="/Team:IIT_Delhi/Results">Squarewave Generator</a>
  
 
       <a href="/Team:IIT_Delhi/Interlab">Interlab</a>
 
       <a href="/Team:IIT_Delhi/Interlab">Interlab</a>
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   </div>  
 
   </div>  
 
                                                         <div class="dropdown">
 
                                                         <div class="dropdown">
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     <div class="dropdown-content">
 
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       <a href="/Team:IIT_Delhi/Basic_Part">Basic Parts</a>
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       <a href="/Team:IIT_Delhi/Basic_Part">Circuit design and construction</a>
−
       <a href="/Team:IIT_Delhi/Composite_Part">Composite Parts</a>
+
       <a href="/Team:IIT_Delhi/Composite_Part">Microfluidics and Fluroscence</a>
−
       <a href="/Team:IIT_Delhi/Improved_Part">Improved Parts</a>
+
       <a href="/Team:IIT_Delhi/Improved_Part">Photobleaching</a>
−
       <a href="/Team:IIT_Delhi/Part_Collection">Part Collection</a>
+
       <a href="/Team:IIT_Delhi/Part_Collection">Promoter strength</a>
 +
                                                      <a href="/Team:IIT_Delhi/Part_Collection">Oscillations</a>
 +
                                                      <a href="/Team:IIT_Delhi/Part_Collection">Parts</a>
 
     </div>
 
     </div>
 
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   </div>
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     <div class="dropdown-content-big">
 
     <div class="dropdown-content-big">
 
       <a href="/Team:IIT_Delhi/Model">Overview</a>
 
       <a href="/Team:IIT_Delhi/Model">Overview</a>
−
       <a href="/Team:IIT_Delhi/Database">Simulation Database</a>
+
       <a href="/Team:IIT_Delhi/Database">Writing a model</a>
 +
                                                        <a href="/Team:IIT_Delhi/Database">Deterministic model </a>
 +
                                                        <a href="/Team:IIT_Delhi/Database">Stochastic model</a>
 +
                                                        <a href="/Team:IIT_Delhi/Database">Bifurcation and squareness</a>
 +
                                                        <a href="/Team:IIT_Delhi/Database">Resource sharing</a>
 
     </div>
 
     </div>
 
     </div>
 
     </div>
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       <a href="/Team:IIT_Delhi/Engagement">Public Engagement</a>
 
       <a href="/Team:IIT_Delhi/Engagement">Public Engagement</a>
 
       <a href="/Team:IIT_Delhi/Collaborations">Collaborations</a>
 
       <a href="/Team:IIT_Delhi/Collaborations">Collaborations</a>
 +
                                                        <a href="/Team:IIT_Delhi/Collaborations">Safety</a>
 
     </div>
 
     </div>
 
   </div>  
 
   </div>  
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−
<!-- Banner -->
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<!--  
 
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CLICK ON THE IMAGES TO KNOW MORE ABOUT US
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<center><img id="team" src="https://static.igem.org/mediawiki/2017/d/da/T--IIT_Delhi--squad.jpg" width = "1300" alt="" ></center>
 
 
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            <h2 class="h2font">LAB PROTOCOLS</h2>
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</div>
  
−
            <p> &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
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</p>
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<h2 id="pfont">Accurate Experimenting for Accurate Results !!</h2>
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          </header>
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          <button class="accordion back1" style="font-weight: bold;">Transformation</button>
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          <div class="panel ">
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<!--<u><b><h4 id="pfont" style="font-size: 90%;">Preparation of competent  cells:-</h4></b></u>-->
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<b id="pfont">Preparation of competent  cells:-</b>
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<div align="left" style="font-size: 84%;">
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<ol ><left>
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−
<li>Dilute an overnight culture of E. coli 1:200 with LB broth.
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<li>Incubate at 37°C with shaking (at 200 rpm) until the cells reach early log phase (OD600 = 0.25-0.4).
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<li>We already have 1X TSS in 4 deg fridge(old). Use it without dilution or thawing. Keep it inside the icebox just after taking out from the fridge. <br>
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−
OR <br>
+
−
(if the above 1X TSS is not available)<br>
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−
While cells are growing, thaw 2X TSS on ice and dilute an appropriate amount 1:1 with sterile distilled water (100µl of diluted TSS will be needed for each ml of cells). Chill on ice.  
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−
<li>  Place 2-ml aliquots of early log-phase cells into sterile 2-ml micro-centrifuge tubes and pellet the cells by centrifugation at 4°C  at 3000g for 10 min.(6-8 mins for taking part from Igem kit)
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−
<li> Remove the supernatant and discard. Add 0.2 ml of the ice-cold 1X TSS and place the tubes on ice.
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−
<li>Gently suspend the cells by pipetting.
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<li> Proceed with the transformation protocol below (Step 2), or immediately freeze cells by immersion in liquid nitrogen or a dry ice/ethanol bath. Store the frozen cells at –70°C.
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−
</left>
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−
</ol>
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</div>
 
</div>
  
−
<b id="pfont">Transformation:-</b>
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<ol ><left>
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    <img src= "https://static.igem.org/mediawiki/2017/a/a8/T--IIT_Delhi--Sashi.jpg" height ="500">
−
<li>Thaw frozen TSS-competent cells slowly on ice(if stored at -70°C).
+
  </div>
−
<li>Add 100 pg -200 ng (2.5 to 4 ul)(15ul for ligation product)of DNA to each tube of competent cells. <br>Note:Addition of more than 10ng of DNA may significantly decrease transformation efficiencies.
+
</div>
−
<li>Flick the tubes to mix the cells and DNA and incubate the cells on ice for 30 minutes. <br>
+
  
−
<li> Transfer the tubes to water bath/dry bath(42°C) for 90 seconds.
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<li> Transfer the tubes to ice and incubate for an additional 10 minutes.  
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<li>Add 800 ul (total 1 mL)of LB broth and incubate the cells at 37°C for up to 1 hour with shaking (at 200 rpm).
+
  <div class="overlay-content ">
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    <img src= "https://static.igem.org/mediawiki/2017/a/a8/T--IIT_Delhi--Abhilash.pptx.jpg" height ="500">
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</div>
  
−
<li> Centrifuge the cells at 3000g for  ~ 6min (10 mins after ligation)at 4deg(in temperature control centrifuge).
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   <li> Aspirate the tubes to leave the pellets with 1/4 broth .(keep ~300ul)
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    <li>Plate the cells on-to the appropriate selective or differential medium and incubate overnight at 37°C.Check the procedure for antibiotic.
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  <div class="overlay-content ">
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      <ol> <li>For Ampicillin: 12ul Amp + 188 ul MQ. In MCT spread it on the culture plate before adding the DNA.
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        <li>For Chloramphenicol: 1:1000 volume ratio of antibiotic : culture broth. Directly suspend into the culture broth and spread it on the plate.
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          <li> For Kanamycin: 1:1000 volume ratio of antibiotic : culture broth. Directly suspend into the culture broth and spread it on the plate.
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          </ol>
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          <ul> DNA should be added as soon as the last trace of ice in the tube disappears.
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<ul>Incubate on ice for 30 minutes. Expect a 2-fold loss in TE for every 10 minutes you shorten this step.
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           <script>
 
           <script>

Latest revision as of 16:05, 1 November 2017

iGEM IIT Delhi

Overview Squarewave Generator Interlab
Circuit design and construction Microfluidics and Fluroscence Photobleaching Promoter strength Oscillations Parts
Overview Writing a model Deterministic model Stochastic model Bifurcation and squareness Resource sharing
Human Practices Integrated Practices Public Engagement Collaborations Safety
Lab Records Protocols Recipes Medal Criteria
The Team Attributions
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Kshitij Pratyush Saksham Divya Siddhesh Pulkit Sashi Shreya Zode Nipun Tanuj Ishank Preeti Abhishek Varuni Tarun Abhilash
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Contact Us Address

E-mail: iitd.igem@gmail.com
Undergraduate Laboratory
Department of Biotechnology and Biochemical Engineering, IIT Delhi