Difference between revisions of "Team:CCU Taiwan/Notebook"

 
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<body>
 
<body>
 
   <header>
 
   <header>
         <nav class="navbar navbar-default navbar-inverse navbar-fixed-top">
+
         <nav>
 
             <div class="container">
 
             <div class="container">
 
                 <ul class="front">
 
                 <ul class="front">
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'https://2018.igem.org/Team:CCU_Taiwan';"> <img src="https://static.igem.org/mediawiki/2018/0/08/T--CCU_Taiwan--home_button.png"></img></div>
 
'https://2018.igem.org/Team:CCU_Taiwan';"> <img src="https://static.igem.org/mediawiki/2018/0/08/T--CCU_Taiwan--home_button.png"></img></div>
  
                     <li class="title" style="cursor:pointer;" id="Home"><img class="img_title" src="https://static.igem.org/mediawiki/2018/2/24/T--CCU_Taiwan--aboutus.png"></img><a>About Us</a><hr>
+
                     <li class="title" style="cursor:pointer;" id="Home"><img class="img_title" src="https://static.igem.org/mediawiki/2018/2/24/T--CCU_Taiwan--aboutus.png"></img><a>About Us</a>
 
                         <ul class="sub" id="sub_home" style="cursor:default;">
 
                         <ul class="sub" id="sub_home" style="cursor:default;">
                             <li class="list" id="home1"><a href="https://2018.igem.org/Team:CCU_Taiwan/Team">Team</a></li>
+
                             <a href="https://2018.igem.org/Team:CCU_Taiwan/Team"><li class="list" id="home1">Team</li></a>
<li class="list" id="home2"><a href="https://2018.igem.org/Team:CCU_Taiwan/Attributions">Attributions</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Attributions"><li class="list" id="home2">Attributions</li></a>
<li class="list" id="home2"><a href="https://2018.igem.org/Team:CCU_Taiwan/Medal">Medal</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Medal"><li class="list" id="home3">Medals</li></a>
<li class="list" id="home3"><a href="https://2018.igem.org/Team:CCU_Taiwan/Judge">For Judge</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Judge"><li class="list" id="home4">For Judges</li></a>
<hr>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Achievements"><li class="list" id="home5">Achievements</li></a>
<div id="home_img">
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<img src="https://static.igem.org/mediawiki/2018/5/57/T--CCU_Taiwan--home.png"></img>
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</div>
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<div id="home_word">
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<a>There's a home for us, always.</a>
+
</div>
+
 
                         </ul>
 
                         </ul>
 
                     </li>
 
                     </li>
                     <li class="title" style="cursor:pointer;" id="Project"><img class="img_title" src="https://static.igem.org/mediawiki/2018/6/6f/T--CCU_Taiwan--project.png"></img><a>Project</a><hr>
+
                     <li class="title" style="cursor:pointer;" id="Project"><img class="img_title" src="https://static.igem.org/mediawiki/2018/6/6f/T--CCU_Taiwan--project.png"></img><a>Project</a>
 
<ul class="sub" id="sub_project" style="cursor:default;">
 
<ul class="sub" id="sub_project" style="cursor:default;">
    <li class="list" id="project1"><a href="https://2018.igem.org/Team:CCU_Taiwan/Description">Description</a></li>
+
    <a href="https://2018.igem.org/Team:CCU_Taiwan/Description"><li class="list" id="project1">Description</li></a>
<li class="list" id="project2"><a href="https://2018.igem.org/Team:CCU_Taiwan/Design">Design</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Applied_Design"><li class="list" id="project2">Design</li></a>
<li class="list" id="project3"><a href="https://2018.igem.org/Team:CCU_Taiwan/Results">Results</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Results"><li class="list" id="project3">Results</li></a>
<li class="list" id="project6"><a href="https://2018.igem.org/Team:CCU_Taiwan/Demonstrate">Demonstration</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Demonstrate"><li class="list" id="project4">Demonstration</li></a>
<li class="list" id="project5"><a href="https://2018.igem.org/Team:CCU_Taiwan/InterLab ">InterLab</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/InterLab"><li class="list" id="project5">InterLab</li></a>
<hr>
+
 
</ul>
 
</ul>
 
</li>
 
</li>
                     <li class="title" style="cursor:pointer;" id="Parts"><img class="img_title" src="https://static.igem.org/mediawiki/2018/1/17/T--CCU_Taiwan--part.png"></img><a>Parts</a><hr>
+
                     <li class="title" style="cursor:pointer;" id="Parts"><img class="img_title" src="https://static.igem.org/mediawiki/2018/1/17/T--CCU_Taiwan--part.png"></img><a>Parts</a>
 
                         <ul class="sub" id="sub_parts" style="cursor:default;">
 
                         <ul class="sub" id="sub_parts" style="cursor:default;">
    <li class="list" id="parts1"><a href="https://2018.igem.org/Team:CCU_Taiwan/Parts">Overview</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Parts"><li class="list" id="parts1">Overview</li></a>
<li class="list" id="parts2"><a href="https://2018.igem.org/Team:CCU_Taiwan/Basic_Part">Basic Part</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Basic_Part"><li class="list" id="parts1">Basic Part</li></a>
<li class="list" id="parts3"><a href="https://2018.igem.org/Team:CCU_Taiwan/Composite_Part">Composite Part</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Composite_Part"><li class="list" id="parts2">Composite Part</li></a>
<li class="list" id="parts4"><a href="https://2018.igem.org/Team:CCU_Taiwan/Improve">Improved Part</a></li>
+
<hr>
+
 
                         </ul>
 
                         </ul>
 
                     </li>
 
                     </li>
                     <li class="title" style="cursor:pointer;" id="Modeling"><img class="img_title" src="https://static.igem.org/mediawiki/2018/0/09/T--CCU_Taiwan--model.png"></img><a>Modeling</a><hr>
+
                     <li class="title" style="cursor:pointer;" id="Modeling"><img class="img_title" src="https://static.igem.org/mediawiki/2018/0/09/T--CCU_Taiwan--model.png"></img><a>Modeling</a>
 
                         <ul class="sub" id="sub_modeling" style="cursor:default;">
 
                         <ul class="sub" id="sub_modeling" style="cursor:default;">
                             <li class="list" id="model1"><a href="https://2018.igem.org/Team:CCU_Taiwan/Model">Overview</a></li>
+
                             <a href="https://2018.igem.org/Team:CCU_Taiwan/Model"><li class="list" id="model1">Overview</li></a>
                            <li class="list" id="model2"><a href="https://2018.igem.org/Team:CCU_Taiwan/Enzyme">Enzyme Model</a></li>
+
                            <a href="https://2018.igem.org/Team:CCU_Taiwan/Binding"><li class="list" id="model2">Binding Model</li></a>
<li class="list" id="model3"><a href="https://2018.igem.org/Team:CCU_Taiwan/Lignin">Lignin Model</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Polymer"><li class="list" id="model3">Polymer Model</li></a>
<li class="list" id="model4"><a href="https://2018.igem.org/Team:CCU_Taiwan/Tools">Tools</a></li>
+
<hr>
+
 
                         </ul>
 
                         </ul>
 
                     </li>
 
                     </li>
                     <li class="title" style="cursor:pointer;" id="Drylab"><img class="img_title" src="https://static.igem.org/mediawiki/2018/f/fc/T--CCU_Taiwan--Dry_lab.png"></img><a>Drylab</a><hr>
+
                     <li class="title" style="cursor:pointer;" id="Drylab"><img class="img_title" src="https://static.igem.org/mediawiki/2018/f/fc/T--CCU_Taiwan--Dry_lab.png"></img><a>Product</a>
 
                         <ul class="sub" id="sub_drylab" style="cursor:default;">
 
                         <ul class="sub" id="sub_drylab" style="cursor:default;">
                             <li class="list" id="drylab1"><a href="https://2018.igem.org/Team:CCU_Taiwan/Our_Plan">Overview</a></li>
+
                             <a href="https://2018.igem.org/Team:CCU_Taiwan/Our_Plan"><li class="list" id="drylab1">Analysis</li></a>
<li class="list" id="drylab2"><a href="https://2018.igem.org/Team:CCU_Taiwan/Engineering">Engineering</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Engineering"><li class="list" id="drylab2">Production Line</li></a>
<li class="list" id="drylab3"><a href="https://2018.igem.org/Team:CCU_Taiwan/Safety">Safety</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Safety"><li class="list" id="drylab3">Safety</li></a>
<hr>
+
 
                         </ul>
 
                         </ul>
 
                     </li>
 
                     </li>
    <li class="title" style="cursor:pointer;" id="Human_Practice"><img class="img_title" src="https://static.igem.org/mediawiki/2018/9/96/T--CCU_Taiwan--humanpractice.png"></img><a>HP</a><hr>
+
    <li class="title" style="cursor:pointer;" id="Human_Practice"><img class="img_title" src="https://static.igem.org/mediawiki/2018/9/96/T--CCU_Taiwan--humanpractice.png"></img><a>HP</a>
 
                         <ul class="sub" id="sub_human_practice" style="cursor:default;">
 
                         <ul class="sub" id="sub_human_practice" style="cursor:default;">
                             <li class="list" id="human_practice1"><a href="https://2018.igem.org/Team:CCU_Taiwan/Human_Practices">Overview</a></li>
+
                             <a href="https://2018.igem.org/Team:CCU_Taiwan/Human_Practices"><li class="list" id="human_practice1">Human Practice</li></a>
<li class="list" id="human_practice2"><a href="https://2018.igem.org/Team:CCU_Taiwan/Engagement">Public</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Public_Engagement"><li class="list" id="human_practice2">Public Engagement</li></a>
<li class="list" id="human_practice3"><a href="https://2018.igem.org/Team:CCU_Taiwan/Entrepreneurship">Entrepreneurship</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Entrepreneurship"><li class="list" id="human_practice3">Entrepreneurship</li></a>
<li class="list" id="human_practice4"><a href="https://2018.igem.org/Team:CCU_Taiwan/Intergrate">Intergrate</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/engaging_experts"><li class="list" id="human_practice4">Engaging Experts</li></a>
<hr>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Integrate"><li class="list" id="human_practice5">Integrated HP</li></a>
 
                         </ul>
 
                         </ul>
 
                     </li>
 
                     </li>
 
    <li class="title" style="cursor:pointer;" id="Notebook"><img class="img_title" src="https://static.igem.org/mediawiki/2018/c/c9/T--CCU_Taiwan--notebook.png"></img><a>Notebook</a>
 
    <li class="title" style="cursor:pointer;" id="Notebook"><img class="img_title" src="https://static.igem.org/mediawiki/2018/c/c9/T--CCU_Taiwan--notebook.png"></img><a>Notebook</a>
 
                         <ul class="sub" id="sub_notebook" style="cursor:default;">
 
                         <ul class="sub" id="sub_notebook" style="cursor:default;">
                             <li class="list" id="notebook1"><a href="https://2018.igem.org/Team:CCU_Taiwan/Notebook">Overview</a></li>
+
                             <a href="https://2018.igem.org/Team:CCU_Taiwan/Notebook"><li class="list" id="notebook1">Overview</li></a>
<li class="list" id="notebook2"><a href="https://2018.igem.org/Team:CCU_Taiwan/Collaborations">Collaborations</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Collaborations"><li class="list" id="notebook2">Collaborations</li></a>
<li class="list" id="notebook3"><a href="https://2018.igem.org/Team:CCU_Taiwan/Experiments">Experiments</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Protocols"><li class="list" id="notebook3">Protocols</li></a>
<li class="list" id="notebook4"><a href="https://2018.igem.org/Team:CCU_Taiwan/DATABASE">DATABASE</a></li>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Experiments"><li class="list" id="notebook4">Experiments</li></a>
<hr>
+
<a href="https://2018.igem.org/Team:CCU_Taiwan/Materials"><li class="list" id="notebook5">Materials</li></a>
 
                         </ul>
 
                         </ul>
 
                     </li>
 
                     </li>
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             </div>
 
             </div>
 
         </nav>
 
         </nav>
<!--<div class="scroll-line"></div>
+
 
<div class="scroll-line-bg"></div>
+
<div class="Quick-navigation">
+
  <a href="#A" class="Quick-navigation-item">____Background</a>
+
  <a href="#B" class="Quick-navigation-item">____Calibration 1</a>
+
  <a href="#C" class="Quick-navigation-item">____Calibration 2</a>
+
  <a href="#D" class="Quick-navigation-item">____Calibration 3</a>
+
  <a href="#E" class="Quick-navigation-item">____Calibration 4</a>
+
  <a href="#F" class="Quick-navigation-item">____Calibration 5</a>
+
  <div class="Scroll-progress-indicator"></div>
+
</div>-->
+
 
     </header>
 
     </header>
<div class="background">
+
<div class="indicator">
<div class="bcg"></div>
+
 
 +
<div class="pointerNotebook" id="1"><a href="#ca1">Notebook</a></div>
 +
</div>
 +
 
 +
<div class="backgroundNotebook">
 +
<div class="photoNotebook"><h1 class="bigtitle">NOTEBOOK<h1></div>
 
       <div class="content">
 
       <div class="content">
 +
<p class="first" id="ca1">Notebook</p>
 +
<p class="second">February</p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;Molecular biology class<br>
 +
    &emsp;&emsp;&emsp;&emsp;<I>E. coli</I> genomic DNA preparation<br>
 +
    &emsp;&emsp;&emsp;&emsp;<I>E. coli</I> transformation<br>
 +
</p>
 +
<p class="second">March</p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;Instrument operation</p>
 +
<p class="second">April</p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;Project brainstorming: Product Positioning, Monolignol proportion</p>
 +
<p class="second">May</p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;Culture selection: compare Yeast, <I>E. coli</I>, Acetobacter aceti<br>
 +
&emsp;&emsp;&emsp;&emsp;Gene design
 +
</p>
 +
<p class="second">June</p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;Interlab experiment: Calibration 1, 2, 3</p>
 +
<p class="second">July</p>
 +
<p class="description">&emsp;&emsp;<strong>7/3</strong><br>
 +
&emsp;&emsp;&emsp;&emsp;Start Interlab experiment: cell measurement<br>
 +
&emsp;&emsp;<strong>7/5</strong><br>
 +
&emsp;&emsp;&emsp;&emsp;Yeast Extract–Peptone–Dextrose (YPD) formulation <br>
 +
&emsp;&emsp;<strong>7/9</strong><br>
 +
&emsp;&emsp;&emsp;&emsp;Yeast (X33) culture<br>
 +
&emsp;&emsp;<strong>7/13</strong><br>
 +
    &emsp;&emsp;&emsp;&emsp;Fundraising briefing session<br>
 +
&emsp;&emsp;<strong>7/16</strong><br>
 +
    &emsp;&emsp;&emsp;&emsp;Communicate with NCKU (Interlab & project)<br>
 +
&emsp;&emsp;<strong>7/18</strong><br>
 +
    &emsp;&emsp;&emsp;&emsp;Communicate with BIT (project)<br>
 +
</p>
 +
<p class="second">August</p>
 +
<p class="description"><strong>8/5~8/12</strong></p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;Cell growth, sampling, and assay<br>
 +
&emsp;&emsp;&emsp;&emsp;Digest pGAPZ A(X3)/ pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI, EcoRI<br>
 +
&emsp;&emsp;&emsp;&emsp;Transformation pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18 (15µl) into 20µl ECOS™<br>
 +
&emsp;&emsp;&emsp;&emsp;101 Competent Cells [DH5a]<br>
 +
&emsp;&emsp;&emsp;&emsp;Ligation pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18<br>
 +
&emsp;&emsp;&emsp;&emsp;Digest pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI, EcoRI<br>
 +
</p>
 +
<p class="description"><strong>8/13~8/19</strong></p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;Miniprep pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18<br>
 +
&emsp;&emsp;&emsp;&emsp;Digest pGAPZ A(X3)/ pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI, EcoRI<br>
 +
&emsp;&emsp;&emsp;&emsp;Digest pGAPZ A with SalI<br>
 +
&emsp;&emsp;&emsp;&emsp;Incubate with DNA Polymerase I, Large (Klenow) Fragment remove SalI restriction site<br>
 +
&emsp;&emsp;&emsp;&emsp;Ligation pGAPZ A-SalI(pGAPZ A-1)<br>
 +
&emsp;&emsp;&emsp;&emsp;Transformation of pGAPZ A-SalI(pGAPZ A-1)<br>
 +
&emsp;&emsp;&emsp;&emsp;Minipreparation of pGAPZ A-SalI(pGAPZ A-1)
 +
</p>
 +
<p class="description"><strong>8/20~8/26</strong></p>
 +
<p class="description">&emsp;&emsp;Gel extract<br>
 +
&emsp;&emsp;&emsp;&emsp;Digestion – pGAPZ A-1+HIS4 cut with AgeI and EcoRI (x3)<br>
 +
&emsp;&emsp;&emsp;&emsp;Ligation - pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18
 +
</p>
 +
<p class="description"><strong>8/27~9/2</strong></p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;Digestion – pGAPZ A-1+HIS4 cut with AgeI and EcoRI (x3)<br>
 +
&emsp;&emsp;&emsp;&emsp;Ligation – pGAPZ A-1+HIS4 cut with AgeI and EcoRI (x3)<br><br>
  
            <p class="first">Notebook</p>
+
&emsp;&emsp;&emsp;&emsp;Transform <br>
<p class="second">February</p>
+
&emsp;&emsp;&emsp;&emsp;&emsp;&emsp;1. pGAPZ A-1 + HIS4 + Px16 <br>
<p class="description">Molecular biology class<br>E-coli genomic DNA preparation<br>E-coli transformation</p>
+
&emsp;&emsp;&emsp;&emsp;&emsp;&emsp;2. pGAPZ A-1 + HIS4 + Px18<br>
<p class="second">March</p>
+
&emsp;&emsp;&emsp;&emsp;&emsp;&emsp;3. pGAPZ A-1 + HIS4 + Lac1<br>
<p class="description">Instrument operation</p>
+
&emsp;&emsp;&emsp;&emsp;Digest pGAPZ A HIS4(X2) with AgeI, EcoRI<br>
<p class="second">April</p>
+
&emsp;&emsp;&emsp;&emsp;pGAPZ A_Px16, pGAPZ A_Px18 , pGAPZ A_Lac1 Traditional plasmid DNA extraction<br>
<p class="description">Project brainstorming-Product Positioning, HGS monomer proportion</p>
+
&emsp;&emsp;&emsp;&emsp;Yeast SMD1168 Transformation(pGAPZ A_Px16, pGAPZ A_Px18 , pGAPZ A_Lac1)
<p class="second">May</p>
+
</p>
<p class="description">Culture selection-compare Yeast, E-coli, Acetobacter aceti<br>Gene design</p>
+
<p class="second">June</p>
+
<p class="description">Interlab experiment-Calbration1,2,3</p>
+
<p class="second">July</p>
+
<p class="description">7/3<br>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;Start interlab experiment-cell measurement<br>7/5<br>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;YPD formulation<br>7/9<br>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;Yeast(X33) culture<br>7/13<br>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;Fundraising briefing session<br>7/16<br>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;Communicate with NCKU(interlab &
+
project)<br>7/18<br>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;Communicate with BIT(project)</p>
+
<p class="description">8/5<br>19:00-20:00<br>&nbsp;&nbsp;Day 2:<br>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;Pick 2 colonies from each of the transformation plates and inoculate in 5mL LB medium +Chloramphenicol. Grow the cells overnight (14 hours) at 37°C and 220 rpm.</p>
+
<p class="description">8/6<br>10:00-18:00<br>&nbsp;&nbsp;Day 3: Cell growth, sampling, and assay<br>
+
<ol>
+
<li>Make a 1:10 dilution of each overnight culture in LB+Chloramphenicol (0.5mL of culture into 4.5mL of LB+Chlor)</li>
+
<li>Measure Abs600 of these 1:10 diluted cultures</li>
+
<li>Record the data in your notebook</li>
+
<li>Dilute the cultures further to a target Abs600 of 0.02 in a final volume of 12 ml LB medium + Chloramphenicol in 50 mL falcon tube (amber, or covered with foil to block light).</li>
+
<li>Take 500 μL samples of the diluted cultures at 0 hours into 1.5 ml eppendorf tubes, prior to incubation. (At each time point 0 hours and 6 hours, you will take a sample from each of the 8 devices, two colonies per device, for a total of 16 eppendorf tubes with 500 μL samples per time point, 32 samples total). Place the samples on ice.</li>
+
<li>Measure your samples (Abs600 and Fluorescence measurement)</li>
+
</ol>
+
</p>
+
<br><br>
+
<p class="description">Digest pGAPZ A(X3)/ pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI,EcoRI</P>
+
<br><br>
+
<p class="description">8/7<br>10:00-12:00<br>Fluorescence measurement</p>
+
<p class="description">Ligation</p>
+
<p class="description">8/8<br>&nbsp;&nbsp;Day 2:<br> &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;Pick 2 colonies from each of the transformation plates and inoculate in 5mL LB medium +Chloramphenicol. Grow the cells overnight (16 hours) at 37°C and 220 rpm.<br>Transformation
+
<ol>
+
<li>Add all pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18 (15µl) into 20µl ECOS™ 101 Competent Cells [DH5a]</li>
+
<li>Incubate on ice 5 minutes</li>
+
<li>Heat shock at 42℃ 45 second</li>
+
<li>Incubate on ice 5 minutes</li>
+
<li>Add 140µl LB</li>
+
<li>Incubate at 37℃ 1hr</li>
+
<li>Spread on LB+ Zeocin plate</li>
+
</ol>
+
</p>
+
<p class="description">Ligation</p>
+
  
 +
<p class="second">September</p>
 +
<p class="description"><strong>9/3~9/9</strong></p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;PCR—pGAPZ A-1 + HIS4 + Px16, Px18, Lac1<br>
 +
&emsp;&emsp;&emsp;&emsp;miniprep – pGAPZ A-1 + HIS4 + Px16, Px18<br>
 +
&emsp;&emsp;&emsp;&emsp;Transform – pGAPZ A-1 + HIS4 + Lac1<br>
 +
&emsp;&emsp;&emsp;&emsp;Digestion – pGAPZ A-1 + HIS4+lac1<br>
 +
</p>
 +
<p class="description"><strong>9/12~9/14</strong></p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;Colony PCR gel electrophoresis check - HIS4+Px16, HIS4+Lac1<br>
 +
&emsp;&emsp;&emsp;&emsp;Culture HIS4+Px16, HIS4+Lac1<br>
 +
&emsp;&emsp;&emsp;&emsp;Colony PCR gel electrophoresis check - HIS4+Px16, HIS4+Lac1<br>
 +
&emsp;&emsp;&emsp;&emsp;Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 colony PCR  <br>
 +
&emsp;&emsp;&emsp;&emsp;Plasmid miniprep - HIS4+Lac1, HIS4+Px16
 +
</p>
 +
<p class="description"><strong>9/24~9/27</strong></p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;Culture HIS4+Lac1 x3<br>
 +
&emsp;&emsp;&emsp;&emsp;Prepare yeast competent cells (SMD1168)<br>
 +
&emsp;&emsp;&emsp;&emsp;Traditional plasmid DNA extraction Day1 <br>
 +
&emsp;&emsp;&emsp;&emsp;Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 colony PCR  <br>
 +
&emsp;&emsp;&emsp;&emsp;Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 Western blot  <br>
 +
&emsp;&emsp;&emsp;&emsp;Prepare YPD plates<br>
 +
&emsp;&emsp;&emsp;&emsp;Prepare the amino acids of SD plates<br>
 +
&emsp;&emsp;&emsp;&emsp;Traditional plasmid DNA extraction Day2<br>
 +
&emsp;&emsp;&emsp;&emsp;Plasmids contain HIS4 cut with SalI<br>
 +
&emsp;&emsp;&emsp;&emsp;Yeast transformation - Px16, Px18, Lac1 (plasmids contain HIS4)
 +
</p>
 +
 +
<p class="second">October</p>
 +
<p class="description"><strong>10/8~10/11</strong></p>
 +
<p class="description">&emsp;&emsp;&emsp;&emsp;Prepare yeast competent cells (SMD1168)<br>
 +
&emsp;&emsp;&emsp;&emsp;Colony PCR (HIS4+Px16)<br>
 +
&emsp;&emsp;&emsp;&emsp;Safety (30℃, 50℃, 70℃)<br>
 +
&emsp;&emsp;&emsp;&emsp;Yeast transformation - Px16, Lac1 (plasmids contain HIS4)<br><br>
 +
&emsp;&emsp;&emsp;&emsp;Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 Western blot  <br>
 +
<br><br>
 +
</p>
 
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Latest revision as of 08:52, 1 December 2018

NOTEBOOK

Notebook

February

    Molecular biology class
    E. coli genomic DNA preparation
    E. coli transformation

March

    Instrument operation

April

    Project brainstorming: Product Positioning, Monolignol proportion

May

    Culture selection: compare Yeast, E. coli, Acetobacter aceti
    Gene design

June

    Interlab experiment: Calibration 1, 2, 3

July

  7/3
    Start Interlab experiment: cell measurement
  7/5
    Yeast Extract–Peptone–Dextrose (YPD) formulation
  7/9
    Yeast (X33) culture
  7/13
    Fundraising briefing session
  7/16
    Communicate with NCKU (Interlab & project)
  7/18
    Communicate with BIT (project)

August

8/5~8/12

    Cell growth, sampling, and assay
    Digest pGAPZ A(X3)/ pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI, EcoRI
    Transformation pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18 (15µl) into 20µl ECOS™
    101 Competent Cells [DH5a]
    Ligation pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18
    Digest pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI, EcoRI

8/13~8/19

    Miniprep pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18
    Digest pGAPZ A(X3)/ pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI, EcoRI
    Digest pGAPZ A with SalI
    Incubate with DNA Polymerase I, Large (Klenow) Fragment remove SalI restriction site
    Ligation pGAPZ A-SalI(pGAPZ A-1)
    Transformation of pGAPZ A-SalI(pGAPZ A-1)
    Minipreparation of pGAPZ A-SalI(pGAPZ A-1)

8/20~8/26

  Gel extract
    Digestion – pGAPZ A-1+HIS4 cut with AgeI and EcoRI (x3)
    Ligation - pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18

8/27~9/2

    Digestion – pGAPZ A-1+HIS4 cut with AgeI and EcoRI (x3)
    Ligation – pGAPZ A-1+HIS4 cut with AgeI and EcoRI (x3)

    Transform
      1. pGAPZ A-1 + HIS4 + Px16
      2. pGAPZ A-1 + HIS4 + Px18
      3. pGAPZ A-1 + HIS4 + Lac1
    Digest pGAPZ A HIS4(X2) with AgeI, EcoRI
    pGAPZ A_Px16, pGAPZ A_Px18 , pGAPZ A_Lac1 Traditional plasmid DNA extraction
    Yeast SMD1168 Transformation(pGAPZ A_Px16, pGAPZ A_Px18 , pGAPZ A_Lac1)

September

9/3~9/9

    PCR—pGAPZ A-1 + HIS4 + Px16, Px18, Lac1
    miniprep – pGAPZ A-1 + HIS4 + Px16, Px18
    Transform – pGAPZ A-1 + HIS4 + Lac1
    Digestion – pGAPZ A-1 + HIS4+lac1

9/12~9/14

    Colony PCR gel electrophoresis check - HIS4+Px16, HIS4+Lac1
    Culture HIS4+Px16, HIS4+Lac1
    Colony PCR gel electrophoresis check - HIS4+Px16, HIS4+Lac1
    Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 colony PCR
    Plasmid miniprep - HIS4+Lac1, HIS4+Px16

9/24~9/27

    Culture HIS4+Lac1 x3
    Prepare yeast competent cells (SMD1168)
    Traditional plasmid DNA extraction Day1
    Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 colony PCR
    Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 Western blot
    Prepare YPD plates
    Prepare the amino acids of SD plates
    Traditional plasmid DNA extraction Day2
    Plasmids contain HIS4 cut with SalI
    Yeast transformation - Px16, Px18, Lac1 (plasmids contain HIS4)

October

10/8~10/11

    Prepare yeast competent cells (SMD1168)
    Colony PCR (HIS4+Px16)
    Safety (30℃, 50℃, 70℃)
    Yeast transformation - Px16, Lac1 (plasmids contain HIS4)

    Yeast SMD1168 pGAPZ A_Px16, pGAPZ A_Px18, pGAPZ A_Lac1 Western blot