Difference between revisions of "Team:Peking/Notebook"

 
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         <meta name="description" content="Wiki of Peking iGEM 2016" />
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         <meta name="description" content="Wiki of Peking iGEM 2018" />
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         <meta name="author" content="Li Jiamian & Wang Yuqing"/>
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         <meta name="author" content="Peking iGEM"/>
 
         <meta http-equiv="Content-Type" content="text/html; charset=utf-8" />
 
         <meta http-equiv="Content-Type" content="text/html; charset=utf-8" />
 
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         <!-- Fix  Overwrite the original iGEM style=================================================== -->
 
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         <link href="https://2016.igem.org/Template:Peking/css/fix?action=raw&ctype=text/css" rel="stylesheet" />
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         <link href="https://2018.igem.org/Template:Peking/css/fix?action=raw&ctype=text/css" rel="stylesheet" />
 
         <!-- CSS======================================================================================= -->
 
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         <!-- CSS======================================================================================= -->
 
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         <link rel="stylesheet" href="https://2016.igem.org/Template:Peking/css/default?action=raw&ctype=text/css"/>
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         <link rel="stylesheet" href="https://2018.igem.org/Template:Peking/css/default?action=raw&ctype=text/css"/>
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         <link rel="stylesheet" href="https://2016.igem.org/Template:Peking/css/layout?action=raw&ctype=text/css"/>
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         <link rel="stylesheet" href="https://2018.igem.org/Template:Peking/css/layout?action=raw&ctype=text/css"/>
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         <link rel="stylesheet" href="https://2016.igem.org/Template:Peking/css/media-queries?action=raw&ctype=text/css"/>
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         <link rel="stylesheet" href="https://2018.igem.org/Template:Peking/css/media-queries?action=raw&ctype=text/css"/>
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         <link rel="stylesheet" href="https://2016.igem.org/Template:Peking/css/notebook_panel?action=raw&ctype=text/css"/>
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         <link rel="stylesheet" href="https://2018.igem.org/Template:Peking/css/notebook_panel?action=raw&ctype=text/css"/>
 
         <style>
 
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                        body { background: #D2D8D8 url(https://static.igem.org/mediawiki/2018/7/78/T--Peking--images_bodyBackground.jpeg); background-attachment:fixed;}
 
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                                     <li><a href="https://2018.igem.org/Team:Peking/Design" class="barfont1">Design</a></li>
 
                                     <li><a href="https://2018.igem.org/Team:Peking/Design" class="barfont1">Design</a></li>
 
                                     <li><a href="https://2018.igem.org/Team:Peking/Demonstrate" class="barfont1">Demonstration</a></li>
 
                                     <li><a href="https://2018.igem.org/Team:Peking/Demonstrate" class="barfont1">Demonstration</a></li>
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                                     <li><a href="https://2018.igem.org/Team:Peking/Prospective" class="barfont1">Prospective</a></li>
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                                     <li><a href="https://2018.igem.org/Team:Peking/Perspective" class="barfont1">Perspective</a></li>
 
                                 </ul>
 
                                 </ul>
 
                             </li>
 
                             </li>
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                             <li class="dropdown menu-3"><a class="dropdown-toggle" data-toggle="dropdown" href="#" >Modeling</a>
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                             <li class="menu-3"><a class="colapse-menu1" href="https://2018.igem.org/Team:Peking/Model">Modeling</a>
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                                <ul class="dropdown-menu">
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−
                                    <li><a href="https://2018.igem.org/Team:Peking/Model">Overview</a></li>
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                                    <li><a href="https://2018.igem.org/Team:Peking/SPOT_Formation" class="barfont1">SPOT Formation</a></li>
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                                    <li><a href="https://2018.igem.org/Team:Peking/Application" class="barfont1">Application</a></li>
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−
                                </ul>
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                             </li>
 
                             </li>
 
                             <li class="menu-4"><a class="colapse-menu1" href="https://2018.igem.org/Team:Peking/Software">Software</a>
 
                             <li class="menu-4"><a class="colapse-menu1" href="https://2018.igem.org/Team:Peking/Software">Software</a>
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                                         <li><a href="https://2018.igem.org/Team:Peking/Collaborations" class="barfont1">Collaborations</a></li>
 
                                         <li><a href="https://2018.igem.org/Team:Peking/Collaborations" class="barfont1">Collaborations</a></li>
 
                                         <li><a href="https://2018.igem.org/Team:Peking/Safety" class="barfont1">Safety</a></li>
 
                                         <li><a href="https://2018.igem.org/Team:Peking/Safety" class="barfont1">Safety</a></li>
−
                                                                            <li><a href="https://2018.igem.org/Team:Peking/Acknowledgement" class="barfont1">Acknowledgement</a></li></ul>
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                                        <li><a href="https://2018.igem.org/Team:Peking/Acknowledgement" class="barfont1">Acknowledgement</a></li>
 +
                                       
 +
                                    </ul>
 
                                 </li>
 
                                 </li>
 
                                 </div>
 
                                 </div>
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<a href="https://static.igem.org/mediawiki/2018/5/57/T--Peking--Carotene_Extraction_Protocol.pdf">Carotene Extraction Protocol.pdf</a><br/>
 
<a href="https://static.igem.org/mediawiki/2018/5/57/T--Peking--Carotene_Extraction_Protocol.pdf">Carotene Extraction Protocol.pdf</a><br/>
  
−
<a href="https://static.igem.org/mediawiki/2018/7/7e/T--Peking--Carotene_Identify_Protocol.pdf">Carotene Identify Protocol.pdf</a><br/>
+
<a href="https://static.igem.org/mediawiki/2018/7/7e/T--Peking--Carotene_Identify_Protocol.pdf">Carotene Identification Protocol.pdf</a><br/>
  
 
<a href="https://static.igem.org/mediawiki/2018/2/24/T--Peking--Carotene_Production_Protocol.pdf">Carotene Production Protocol.pdf</a><br/>
 
<a href="https://static.igem.org/mediawiki/2018/2/24/T--Peking--Carotene_Production_Protocol.pdf">Carotene Production Protocol.pdf</a><br/>
  
−
<a href="https://static.igem.org/mediawiki/2018/c/cd/T--Peking--Molecular_Cloning_%28Plasmid_extraction_DNA_electrophoresis_DNA_clean%29.pdf">Molecular Cloning Plasmid extraction DNA electrophoresis DNA clean.pdf</a><br/>
+
<a href="https://static.igem.org/mediawiki/2018/c/cd/T--Peking--Molecular_Cloning_%28Plasmid_extraction_DNA_electrophoresis_DNA_clean%29.pdf">Molecular Cloning Plasmid extraction/DNA electrophoresis/DNA clean.pdf</a><br/>
  
−
<a href="https://static.igem.org/mediawiki/2018/4/49/T--Peking--Molecular_Cloning_Protocol_%28DNA_recombanation%29.pdf">Molecular Cloning Protocol DNA recombanation.pdf</a><br/>
+
<a href="https://static.igem.org/mediawiki/2018/4/49/T--Peking--Molecular_Cloning_Protocol_%28DNA_recombanation%29.pdf">Molecular Cloning Protocol DNA recombination.pdf</a><br/>
  
 
<a href="https://static.igem.org/mediawiki/2018/9/9d/T--Peking--Molecular_Cloning_Protocol_%28E_coli_transformation%29.pdf">Cloning Protocol E.coli transformation.pdf</a><br/>
 
<a href="https://static.igem.org/mediawiki/2018/9/9d/T--Peking--Molecular_Cloning_Protocol_%28E_coli_transformation%29.pdf">Cloning Protocol E.coli transformation.pdf</a><br/>
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<a href="https://static.igem.org/mediawiki/2018/c/cd/T--Peking--Yeast_Transformation_Protocol.pdf">Yeast Transformation Protocol.pdf</a><br/>
 
<a href="https://static.igem.org/mediawiki/2018/c/cd/T--Peking--Yeast_Transformation_Protocol.pdf">Yeast Transformation Protocol.pdf</a><br/>
 +
 +
<a href="https://static.igem.org/mediawiki/2018/9/96/T--Peking--yeastweighprotocol.pdf">Yeast Dry Weight and OD Measurement Protocol.pdf</a><br/>
 +
  
 
                                 </p></div>
 
                                 </p></div>
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</p>
 
</p>
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                                 <div class="info">
 
                                 <div class="info">
 
                               <div class="ordi">10</div>
 
                               <div class="ordi">10</div>
 
                                 </div>
 
                                 </div>
 
                                 <div class="content">
 
                                 <div class="content">
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                                     <h3>2018.7.16-7.22
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                                     <h3>2018.7.16-2018.7.22
  
 
</h3>
 
</h3>
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                                 <div class="content">
 
                                 <div class="content">
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                                     <p>Completed Interlab tasks.<br/>
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                                     <p>Meeting up with Fudan-CHINA.<br/>
−
Meeting up with Fudan-CHINA.<br/>
+
 
Insert 4 kinds of promoters PDH3, TEF1, Ura3, Tet07 into 4 kinds of yeast transformation plasmids pNH603, pNH604, pNH605, pNH606. <br/>
 
Insert 4 kinds of promoters PDH3, TEF1, Ura3, Tet07 into 4 kinds of yeast transformation plasmids pNH603, pNH604, pNH605, pNH606. <br/>
  
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  <div class="coll">
 
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</p>
 
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<div class="coll">
 
                                 <div class="info">
 
                                 <div class="info">
 
                               <div class="ordi">23</div>
 
                               <div class="ordi">23</div>
 
                                 </div>
 
                                 </div>
 
                                 <div class="content">
 
                                 <div class="content">
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                                     <h3>2018.10.15-17
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                                     <h3>2018.10.15-2018.10.17
 +
 
 
</h3>
 
</h3>
 
                                 </div>
 
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                                 <div class="content">
 
                                     <p>WIKI and poster making.<br/>
 
                                     <p>WIKI and poster making.<br/>
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Presentation preparation.  
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Presentation preparation.
  
  
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         <style>
 
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                     <ul class="footer-social">
 
                     <ul class="footer-social">
 
                         <li class="col-md-6" id="PKU-administration" style="margin-bottom:25px;max-width:300px">
 
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                             <a href="http://dean.pku.edu.cn/pkudean/index.html"><img src="https://static.igem.org/mediawiki/2018/7/7a/T--Peking--images_PKU_Administration_logo.png"></a>
 
                         </li>
 
                         </li>
 
                         <li class="col-md-6" id="PKU-SLS" style="margin-bottom:25px;max-width:300px">
 
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                             <a href="http://www.bio.pku.edu.cn/"><img src="https://static.igem.org/mediawiki/2018/0/04/T--Peking--images_PKU_SLS_logo.png"></a>
 
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                         <li class="col-md-6" id="PKU-CQB" style="margin-bottom:25px;max-width:300px">
 
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                             <a href="http://cqb.pku.edu.cn/en/"><img src="https://static.igem.org/mediawiki/2018/e/e1/T--Peking--images_PKU_CQB_logo.png"></a>
 
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                     </ul>
 
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                     <ul class="copyright">
 
                     <ul class="copyright">
 
                         <!--<li>&copy; 2014 Sparrow</li> -->
 
                         <!--<li>&copy; 2014 Sparrow</li> -->
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                         <li><a href="https://2018.igem.org/Team:Peking">Home</a>&nbsp;&nbsp;&nbsp;<a href="mailto:indigomad@pku.edu.cn">Contact</a></li>
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                         <li><a href="https://2018.igem.org/Team:Peking">Home</a>&nbsp;&nbsp;&nbsp;<a href="mailto:pekingigem2018@126.com">Contact</a></li>
 
                         <span> &copy;2018 PEKING IGEM. All Rights Reserved.</span>
 
                         <span> &copy;2018 PEKING IGEM. All Rights Reserved.</span>
 
                         <li><a href="http://getbootstrap.com/2.3.2/">Based on Bootstrap</a></li>
 
                         <li><a href="http://getbootstrap.com/2.3.2/">Based on Bootstrap</a></li>
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            <!-- Java Script======================================================================= -->
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Latest revision as of 00:37, 18 October 2018

Notebook

Protocol

Carotene Extraction Protocol.pdf
Carotene Identification Protocol.pdf
Carotene Production Protocol.pdf
Molecular Cloning Plasmid extraction/DNA electrophoresis/DNA clean.pdf
Molecular Cloning Protocol DNA recombination.pdf
Cloning Protocol E.coli transformation.pdf
Molecular Cloning Protocol PCR.pdf
Yeast Transformation Protocol.pdf
Yeast Dry Weight and OD Measurement Protocol.pdf

Diary
0

2018.5.13

Confirmed project plan.
Prepared experiment equipment and reagents.

1

2018.5.14-2018.5.20

Molecular cloning test.
Yeast transformation test.

2

2018.5.21-2018.5.27

Modeling beginning.

3

2018.5.28-2018.6.3

Plasmid backbone propagation：
E. coli expression: pET-28a
yeast expression: pNH603, pNH604, pNH605, pNH606
Required gene segment synthesis beginning.

4

2018.6.4-2018.6.10

Gibson assembly, golden gate assembly, and other seamless cloning test.
IPTG induced expression test.

5

2018.6.11-2018.6.17

Restriction recombination test.
Began to construct SUMO-SIM system.

6

2018.6.18-2018.6.24

Construction E. coli expression plasmid：
pET-28a-FKBP-yEGFP-HOTag3
pET-28a-Frb-yEGFP-HOTag6

7

2018.6.25-2018.7.1

Fluorescence observation test.

8

2018.7.2-2018.7.8

Joined iGEM conference at Nankai University held by NKU_CHINA.

9

2018.7.9-2018.7.15

Construction yeast transformation plasmid of carotene synthesis:
pNH603-crtE/crtI/crtYB
pNH604-crtE/crtI/crtYB
pNH605-crtE/crtI/crtYB
pNH606-crtE/crtI/crtYB

10

2018.7.16-2018.7.22

Meeting up with Fudan-CHINA.
Insert 4 kinds of promoters PDH3, TEF1, Ura3, Tet07 into 4 kinds of yeast transformation plasmids pNH603, pNH604, pNH605, pNH606.

11

2018.7.23-2018.7.29

Construction yeast transformation plasmids of SPOT:
pNH604-PDH3/TEF1/Ura3/Tet07-FKBP-yEGFP-HOTag3
pNH605-PDH3/TEF1/Ura3/Tet07-Frb-yEGFP-HOTag6
pNH604-PDH3/TEF1/Ura3/Tet07-SUMO-yEGFP-HOTag3
pNH605-PDH3/TEF1/Ura3/Tet07-SIM-yEGFP-HOTag6

12

2018.7.30-2018.8.5

Transferred FKBP-Frb SPOT system and SUMO-SIM SPOT system into yeast.
Nucleic localization test.
Fluorescence microscope observation and flow cytometry measurement of transformed yeast cell lines.

13

2018.8.6-2018.8.12

Rapamycin induction concentration test of FKBP-Frb SPOT system.

Construction yeast transformation plasmids of carotene production:
pNH603-PDH3/TEF1/Ura3/Tet07-Frb-crtI-HOTag6
pNH605-PDH3/TEF1/Ura3/Tet07-Frb-crtE-HOTag6
pNH606-PDH3/TEF1/Ura3/Tet07-Frb-crtYB-HOTag6

Construction yeast transformation plasmids of light induced SPOT system:

pNH604-PDH3/TEF1/Ura3/Tet07-phyB-yEGFP-HOTag3
pNH605-PDH3/TEF1/Ura3/Tet07-PIF-yEGFP-HOTag6
pNH604-PDH3/TEF1/Ura3/Tet07-VVDn-yEGFP-HOTag3
pNH605-PDH3/TEF1/Ura3/Tet07-VVDp-yEGFP-HOTag6

14

2018.8.13-2018.8.19

Carotene production and extraction test.

Construction yeast transformation plasmids of carotene production with SPOT:
pNH603--PDH3/TEF1/Ura3/Tet07-Frb-crtI-HOTag6
pNH605--PDH3/TEF1/Ura3/Tet07-Frb-crtE-HOTag6
pNH606--PDH3/TEF1/Ura3/Tet07-Frb-crtYB-HOTag6
pNH603--PDH3/TEF1/Ura3/Tet07-SIM-crtI-HOTag6
pNH605--PDH3/TEF1/Ura3/Tet07-SIM-crtE-HOTag6
pNH606--PDH3/TEF1/Ura3/Tet07-SIM-crtYB-HOTag6

15

2018.8.20-2018.8.26

Carotene production and extraction test.

Construction yeast transformation plasmids of carotene production with SPOT:
pNH603--PDH3/TEF1/Ura3/Tet07-Frb-crtI-HOTag6
pNH605--PDH3/TEF1/Ura3/Tet07-Frb-crtE-HOTag6
pNH606--PDH3/TEF1/Ura3/Tet07-Frb-crtYB-HOTag6
pNH603--PDH3/TEF1/Ura3/Tet07-SIM-crtI-HOTag6
pNH605--PDH3/TEF1/Ura3/Tet07-SIM-crtE-HOTag6
pNH606--PDH3/TEF1/Ura3/Tet07-SIM-crtYB-HOTag6

16

2018.8.27-2018.9.2

Carotene production and extraction test.

Construction the control group plasmids of SPOT:

pNH604-PDH3/TEF1/Ura3/Tet07-FKBP-yEGFP
pNH605-PDH3/TEF1/Ura3/Tet07-Frb-yEGFP
pNH604-PDH3/TEF1/Ura3/Tet07-SUMO-yEGFP
pNH605-PDH3/TEF1/Ura3/Tet07-SIM-yEGFP
pNH604-PDH3/TEF1/Ura3/Tet07-FKBP-mCherry
pNH605-PDH3/TEF1/Ura3/Tet07-Frb-mCherry
pNH604-PDH3/TEF1/Ura3/Tet07-SUMO-mCherry
pNH605-PDH3/TEF1/Ura3/Tet07-SIM-mCherry
pNH603-H2A-CFP

Joined the 5th CCiC at ShanghaiTech.

17

2018.9.3-2018.9.9

Growth curve measurement of the SPOT system yeast.
Characterization of the carotene production by SPOT system.

18

2018.9.10-2018.9.16

The ability of phase separation induced by rapamycin in FKBP-Frb SPOT system test and longer time observation.
Yeast fermentation test.

19

2018.9.17-2018.9.23

Completed the two kinds of fluorescence repeat in both FKBP-Frb and SUMO-SIM SPOT system.
Carotene production and quantification test.

Construction yeast transformation plasmids of ABA induced SPOT system:
pNH604-PDH3/TEF1/Ura3/Tet07-ABI-yEGFP-HOTag3
pNH605-PDH3/TEF1/Ura3/Tet07-PYL1-yEGFP-HOTag6

Construction yeast transformation plasmids of GA induced SPOT system:
pNH604-PDH3/TEF1/Ura3/Tet07-GAI-yEGFP-HOTag3
pNH605-PDH3/TEF1/Ura3/Tet07-GID-yEGFP-HOTag6

20

2018.9.24-2018.9.30

FRAP test of FKBP-Frb SPOT system.
BioBrick submitted bioparts construction.

21

2018.10.1-2018.10.7

Construction yeast transformation plasmids of GFP nanobody SPOT system:
pNH605-PDH3/TEF1/Ura3/Tet07-Frb-GFPnanobody-HOTag3

Construction yeast transformation plasmids of NAD sensor SPOT system:
pNH604-PDH3/TEF1/Ura3/Tet07-NAD1A-yEGFP-HOTag3
pNH605-PDH3/TEF1/Ura3/Tet07-NAD1B-yEGFP-HOTag6

22

2018.10.8-2018.10.14

Data collection and modeling processing.
HPLC quantification of carotene.

23

2018.10.15-2018.10.17

WIKI and poster making.
Presentation preparation.