Difference between revisions of "Team:UCAS-China/Contributions"

 
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<h1> Team Accomplishments</h1>
 
  
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<h2>Project:<h2>
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<h2>CONTRIBUTIONS</h2>
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<p>The prototype of our project was raised by Yun DONG and Yiyi WANG in our brainstorm session in Feb. 2018. This idea was then modified and improved by Yuanli GAO. Our investigators Prof. Chunbo LOU and Prof. Jiangyun WANG helped in our project design. Our advisors Fankang MENG also offered help in project design.</p>
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<p>We participated in the InterLab study, which helped to reduce the lab-to-lab variability around the world. We also put forward new ideas of the tandem expression of chromoproteins, to get new colors, which would greatly enrich the colors and the BioBrick parts in the Registry. Then using the RGB system[1] which was central to our project, we characterized the Red-, Green-,Blue light sensors (Cph8, YF1, CcaSR) which had already existed in the registry(BBa_K1886006, BBa_K1053210, BBa_K2598005), by exploring the relationship between the fluorescent intensity with the wavelengths and intensity of input light.  
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<h2>Experiments:<h2>
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<h2>INTERLAB</h2>
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<p>Our experiments started in July 2017. As our team's leader, Yuanli GAO designed and monitored our experiments. He also did the primary work of data analysis. Our advisors Fankang MENG and Xiaohong LIU gave their suggestions on project design. Our experiments were accomplished with the joint effort of the following 9 teammates: Yuanli GAO, Yun Dong, Ziyi ZHAO, Zpeng XU, Yiyi WANG, Liangpu WANG, Yuyan NI, Zhaoying PAN.</p>
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<h4>Summary</h4>
  
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<h2>Hardware:<h2>
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<p>Interlab is a global collaboration through which the Measurement Committee aim to improve the measurement tools available to both the iGEM community and the synthetic biology community as a whole. This year, the Committee along with teams all over the world aimed to reduce lab-to-lab variability in fluorescence measurements by normalizing to absolute cell count or colony-forming units (CFUs) instead of OD.</p>
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<p>Our hardware was designed by Zhaoying PAN, Yuyan NI, while Shiyang WANG participated in the discussion. Zhaoying PAN and Yuyan NI build the prototype device with accessible materials in our lab. Zhaoying PAN designed and made the final hardware.</p>
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<h2>Software:<h2>
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<h4>Procedure</h4>
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<p>Our software was designed by Yun DONG and Shiyang WANG. The software was written by Shiyang WANG.</p>
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<p>In order to compute the cell count in samples, two orthogonal approaches were exploited. (detailed protocol was provided in https://2018.igem.org/Measurement/InterLab/Plate_Reader)</p>
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<h2>Modelling:<h2>
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<p>Our modeling work was mainly accomplished by Liangpu Wang, with the help of Shiyang WANG.</p>
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<h2>Human Practice:<h2>
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<li> Converting between absorbance of cells to absorbance of a known concentration of beads:</li>
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<p>Yun DONG is the leader of our HP Squad. She accomplished most of the contact and interview work and record every HP activity. Jiachen HU and Yaru TIAN made the educational videos. Yufei SHAN contacted the educational speech in the primary school and some meet-ups. Yuanli GAO, Ziyi ZHAO, Zepeng XU, Yiyi WANG, Liangpu WANG, Yuyan NI, Zhaoying PAN also contributed to the HP work.</p>
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<p> In this year’s Measurement Kit, we were provided with a sample containing silica beads that were roughly the same size and shape as a typical E. coli cell. As we knew the concentration of the beads, we could convert absorbance measurements into a universal, standard “equivalent concentration of beads” measurement.
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<li> Counting colony-forming units (CFUs) from the sample:
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<p> Since each colony began with a single cell, by counting the colonies cell numbers in certain volume of media could be determined. We were required to determine the number of CFUs in positive and negative control samples in order to compute a conversion factor from absorbance to CFU.
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<h4>Results<h4>
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<img class="img-responsive img-center" width="600px;" src="https://static.igem.org/mediawiki/2018/a/a2/T--UCAS-China--interlab1.jpg">
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<h5>Fig 1. Particle standard curve of Silica beads for Abs​600  calibration</h5>
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<img class="img-responsive img-center" width="600px;" src="https://static.igem.org/mediawiki/2018/9/91/T--UCAS-China--interlab2.jpg">
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<h5>Fig 2. Fluorescein standard curve for fluorescence calibration</h5>
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<h2>Wiki:<h2>
 
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<p>Our wiki was designed by Shiyang WANG, Yaru TIAN and Yufei SHAN. The coding work was finished by Yuanli GAO, Shiyang WANG, with the help of Fankang Meng. The paperwork was finished by the joint effort of Yuanli GAO, Yun Dong, Ziyi ZHAO, Zpeng XU, Yiyi WANG, Liangpu WANG, Yuyan NI, Zhaoying PAN, and finally summarized and modified by Yuanli GAO.</p>
 
  
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<h2>Art work:<h2>
 
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<p>Our art work was accomplished by our excellent art designer Yaru TIAN, Yufei SHAN, while Shiyang WANG participated in the discussion.</p>
 
  
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<h1>Supports<h1>
 
  
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<h2>General support:<h2>
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<h2>TANDEM EXPRESSION OF CHROMOPROTEINS</h2>
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<p>University of Chinese Academy of Sciences</p>
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<p>We put forward a new concept—mixing color into bacterial cells. Unlike the mix of different bacterial cells which produce different colors as the previous iGEM teams have done, we used tandem expression and RGB system to control the ratios of the expression of different colors in bacterial cells, to achieve mixing color in bacterial cells, and to produce more colors from limited chromoproteins, which would greatly enrich the colors and the BioBrick parts in the Registry. (More information, please see <a href= "https://2018.igem.org/Team:UCAS-China/Validated_contribution">Valid Contribution & Proof of Concept</a>)</p>
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<p>College of Life Sciences, University of Chinese Academy of Sciences</p>
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<img class="img-responsive img-center" width="600px;" src="https://static.igem.org/mediawiki/2018/a/a5/T--UCAS-China--color4.png">
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<p>Institute of Biophysics, Chinese Academy of Sciences</p>
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<h5>Figure 3. The color spectrum built from chromoproteins and their tandem expression products.</h5>
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<p>Institute of Microorganism, Chinese Academy of Sciences</p>
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<h3>Project support and advice<h3>
 
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<p>Our modeling work was mainly accomplished by Liangpu Wang, with the help of Shiyang WANG.</p>
 
  
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<h4>Modelling:<h4>
 
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<p>Our modeling work was mainly accomplished by Liangpu Wang, with the help of Shiyang WANG.</p>
 
  
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<h2>Modelling:<h2>
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<h2>CHARACTERIZATION OF SENSORS</h2>
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<p>Our modeling work was mainly accomplished by Liangpu Wang, with the help of Shiyang WANG.</p>
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<p>We used our RGB system to further characterize the red-, green-, blue-light sensors (BBa_K1886006, BBa_K1053210, BBa_K2598005), which had already existed in the registry, because the expression of red-, green-, blue-fluorescent proteins would change according to the excitation degrees of red-, green, blue-sensors. (More information, please see <a href= "https://2018.igem.org/Team:UCAS-China/LightToColor">LIGHT TO COLOR</a>) </p><br>
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<img class="img-responsive img-center" width="1000px;" src="https://static.igem.org/mediawiki/2018/b/b2/T--UCAS-China--color12.jpg">
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<h5>Figure 4. The flow cytometry results shows the distribution of the cells in fluorescence intensity. BV421 represented the blue-fluorescence intensity, while the FITC-A represented the green-fluorescence intensity and Pe-TxR-A represented the red-fluorescence intensity. The horizontal axis shew the fluorescence intensity, while the vertical axis shew the number of bacteria cells.</h5>
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Latest revision as of 02:33, 18 October 2018

CONTRIBUTIONS

We participated in the InterLab study, which helped to reduce the lab-to-lab variability around the world. We also put forward new ideas of the tandem expression of chromoproteins, to get new colors, which would greatly enrich the colors and the BioBrick parts in the Registry. Then using the RGB system[1] which was central to our project, we characterized the Red-, Green-,Blue light sensors (Cph8, YF1, CcaSR) which had already existed in the registry(BBa_K1886006, BBa_K1053210, BBa_K2598005), by exploring the relationship between the fluorescent intensity with the wavelengths and intensity of input light.

INTERLAB

Summary

Interlab is a global collaboration through which the Measurement Committee aim to improve the measurement tools available to both the iGEM community and the synthetic biology community as a whole. This year, the Committee along with teams all over the world aimed to reduce lab-to-lab variability in fluorescence measurements by normalizing to absolute cell count or colony-forming units (CFUs) instead of OD.

Procedure

In order to compute the cell count in samples, two orthogonal approaches were exploited. (detailed protocol was provided in https://2018.igem.org/Measurement/InterLab/Plate_Reader)

  • Converting between absorbance of cells to absorbance of a known concentration of beads:

    • In this year’s Measurement Kit, we were provided with a sample containing silica beads that were roughly the same size and shape as a typical E. coli cell. As we knew the concentration of the beads, we could convert absorbance measurements into a universal, standard “equivalent concentration of beads” measurement.

  • Counting colony-forming units (CFUs) from the sample:

    • Since each colony began with a single cell, by counting the colonies cell numbers in certain volume of media could be determined. We were required to determine the number of CFUs in positive and negative control samples in order to compute a conversion factor from absorbance to CFU.

Results

Fig 1. Particle standard curve of Silica beads for Abs​600 calibration


Fig 2. Fluorescein standard curve for fluorescence calibration


TANDEM EXPRESSION OF CHROMOPROTEINS

We put forward a new concept—mixing color into bacterial cells. Unlike the mix of different bacterial cells which produce different colors as the previous iGEM teams have done, we used tandem expression and RGB system to control the ratios of the expression of different colors in bacterial cells, to achieve mixing color in bacterial cells, and to produce more colors from limited chromoproteins, which would greatly enrich the colors and the BioBrick parts in the Registry. (More information, please see Valid Contribution & Proof of Concept)

Figure 3. The color spectrum built from chromoproteins and their tandem expression products.


CHARACTERIZATION OF SENSORS

We used our RGB system to further characterize the red-, green-, blue-light sensors (BBa_K1886006, BBa_K1053210, BBa_K2598005), which had already existed in the registry, because the expression of red-, green-, blue-fluorescent proteins would change according to the excitation degrees of red-, green, blue-sensors. (More information, please see LIGHT TO COLOR)


Figure 4. The flow cytometry results shows the distribution of the cells in fluorescence intensity. BV421 represented the blue-fluorescence intensity, while the FITC-A represented the green-fluorescence intensity and Pe-TxR-A represented the red-fluorescence intensity. The horizontal axis shew the fluorescence intensity, while the vertical axis shew the number of bacteria cells.