Team:NC School of Sci Math/Lab Notebook

From 2013hs.igem.org

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<h2><a name="Calibration" style="color:#0008B;">Calibration and Characterization</a></h2>
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<h2><a name="Experimentation" style="color:#0008B;">Experimentation</a></h2>
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<h4><a name="04/27/2012">04/27/2012</a></h4>
<h4><a name="04/27/2012">04/27/2012</a></h4>
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* We want to work with the ADK for the next couple of weeks
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* Jack will attempt to code the ADK so it outputs to a text file
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* Potential preliminary tests
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** Diluted glow sticks
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** Dimmed LED lights
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** pGLO E. coli
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<h4><a name="05/04/2013">05/04/2013</a></h4>
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* The ADK has been coded
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* We have glow sticks, E. coli expressing GFP, and dim LEDs to test
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* Testing this Tuesday
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<h4><a name="05/07/2013">05/07/2013</a></h4>
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4 ml bacterial culture is pelleted by centrifugation at 10.000 rpm for 1 min. Afterwards, the pellet is resuspended in 250 µl buffer P1. 250 µl buffer P2 are added and the culture is gently inverted and incubated for 5 min. 350 µl buffer N3 are added and the suspension is centrifuged at 13.000 rpm for 10 min. The supernatant is subsequently loaded onto a miniprep column and centrifuged for 1 min/max speed. Afterwards the column is washed with buffer PB (500 µl) and buffer PE (750 µl) by loading onto the column and subsequent centrifugation for 1 min/max speed. After a last centrifugation step for drying the column (again 1 min/max speed), 50 µl of water are added in order to dissolve the DNA again. After 1 min incubation, DNA is eluted by centrifugation for 1 min/max speed.
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* Tested the sensing ability of the ADK today
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* Data is output in the form of the file below
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==Lab Notebook==
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To begin our experiment, we needed to create a device that could detect color. We chose to use the Google ADK, an open ended platform that allows users to take control of many sensors that come attached to an Arduino board, including a colorimeter. However, a standard ADK is set up to match any color presented to it, using LED lights. In some of our preliminary tests, the reflection of the light from these LEDs skewed data. Therefore, we modified the code of the ADK, using Google's software developer package, so that it does not turn on its LED lights but instead outputs colorimeter data to a computer. An example of this output is shown below.
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* The pGLO E. coli is the most closely related to our project
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As a foray into our project, we first took some preliminary data using our modified Google ADK and pGLO E. coli expressing Green Fluorescent Protein. We found that we were able to detect a distinct change in color. Some baseline color was present from the UV Light source used for the experiment. The graphs below clearly show that our system is able to detect E. coli expressing GFP.
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[[File:pGLOGraphs.png|800px]]
[[File:pGLOGraphs.png|800px]]
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* Data from LED lights is very similar but allowed us to understand the sensing "logic" of the ADK
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** Showing red, green, or blue light increases only the color being shown
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** Showing yellow light increases both red and green light
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** Can identify which color is being shown only from the data
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Small LED lights were used to test the ADK's ability to detect red, blue, and yellow lights. The results showed the same trends as the GFP test, indicating that we would be able to distinguish between several different colors expressed. In general, red, green, and blue lights show dramatic increases in the intensity of that color, while the other two colors remain about the same. Yellow light however, increase both red and green light, while blue remains at the same level. From these observations, we are able to identify which type of light is shown based only on the data collected.
 
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Revision as of 13:28, 20 June 2013

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2013


Planning and Development

01/11/2013

Brainstorming for project ideas

  • LDL-cholesterol biosensor
  • Ocean salinity regulator (especially for coral reefs)
  • Water contaminant multibiosensor


More research should be done

  • Find related projects that have been completed
  • Obstacles?
  • BioBricks?
  • Presentations in 3-4 weeks

01/18/2013

Discussions of each project

  • LDL-cholesterol biosensor
    • Could be used as a blood test
    • Blood sugar monitor-like device
    • Hard to measure any color change in blood
  • Ocean salinity regulator (especially for coral reefs)
    • Need a lot of bacteria
    • Environmental concerns
    • Bacteria can do well in coral reefs
  • Water contaminant multibiosensor
    • Several Biobrick promoters for common contaminants
    • Usage in sewer tanks
    • Many different colors of fluorescent proteins
    • Could incorporate newer technology for better color-sensing
    • Will look up data on promoter/FP expression

02/01/2013

Presentations

  • All ideas sound promising
  • Meet next week to decide which idea to pursue

02/08/2013

Decision on project

  • LDL-cholesterol idea won't work
    • Costly and hard to detect FPs in blood
  • Multibiosensor is appealing
    • Many applications
    • Expandable with many different FPs
    • Could bring in new technology
  • Ocean salinity regulator is difficult because of the ocean
    • Environmental factors and unknown variables

Multibiosensor chosen

02/15/2013

A few members met with Mr. Jon Davis to discuss potential technology

  • Suggested a Google device called ADK
    • Comes with a colorimeter
    • Can be programmed for many functions
    • School has a few we could try out

02/22/2013

Goals for the immediate future

  • Find Biobricks
  • Set up schedule for lab work
  • Further research into possible uses
  • Preliminary lab experiments


Project Proposal


03/01/2013

Discussion of upcoming project proposal:

  • Introduction, Solution Statement, and Methods
  • Need to find out some background information
  • Should make use of Tinkercell modeling

03/04/2013

Assignments of roles for the upcoming project proposal:

  • Madeline will investigate background information
  • Danny will work on Tinkercell models
  • Jack will write the solution statement and methods sections
  • Synthesize our writing on a Google Doc
  • Try to have individual parts done in about 2 weeks

03/18/2013

The following models have been created

  • We should use these in the proposal, but continue to work on the Tinkercell model

Design.jpg

03/25/2013

Everyone began editing the proposal on a Google Doc today

  • Want to be finished by March 29th
  • Tinkercells will probably not be ready in time
    • Having a lot of trouble getting fluorescent protein levels to zero when they should be
    • Danny will talk to Morgan to see if he can help
  • Edit Edit Edit!

03/27/2013

  • Proposal is coming along well
  • Need to finish the methods, but otherwise complete
  • Take pictures of the ADK and lab work

03/29/2013

  • Proposal is done!
  • Will submit soon

04/06/2013

Time to focus on creating the plasmid

  • Finished Tinkercell model will help a bunch
  • Do we want to think about ordering de novo from a gene synthesis company?
  • Computer modeling is the best approach for now

04/17/2013

  • Danny has a working Tinkercell model for the copper detector!
    • Outputs increased levels of TagRFP as ion concentration increases
  • We should be able to figure out the others soon
  • The lead detector may be especially difficult because the ion binds to a promoter to create a transcription factor

Experimentation


04/27/2012

  • We want to work with the ADK for the next couple of weeks
  • Jack will attempt to code the ADK so it outputs to a text file
  • Potential preliminary tests
    • Diluted glow sticks
    • Dimmed LED lights
    • pGLO E. coli


<a name="05/04/2013">05/04/2013</a>

</html>

  • The ADK has been coded
  • We have glow sticks, E. coli expressing GFP, and dim LEDs to test
  • Testing this Tuesday


<a name="05/07/2013">05/07/2013</a>

</html>

  • Tested the sensing ability of the ADK today
  • Data is output in the form of the file below

Screenshot.jpg

  • The pGLO E. coli is the most closely related to our project

PGLOGraphs.png

  • Data from LED lights is very similar but allowed us to understand the sensing "logic" of the ADK
    • Showing red, green, or blue light increases only the color being shown
    • Showing yellow light increases both red and green light
    • Can identify which color is being shown only from the data
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