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         <p>A challenge of synthetic biology is repeating measurements in different laboratories. For example, fluorescence data is difficult to compare either because it is reported in different units, or because different groups handle raw data differently.
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         <p>To write an overview.  
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          iGEM’s Measurement Committee thus aims to use the InterLab Study to eventually develop absolute units for measurements of green fluorescent protein (GFP) in a plate reader. This will improve the measurement tools of synthetic biologists.
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        <p> This year, the Committee aims to discover if it is possible to reduce lab-to-lab variability in fluorescence measurements by normalizing to absolute cell count or colony-forming units (CFUs) instead of optical density (OD). For this, we were required
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          to measure the cell density of Escherichia coli (<i>E. coli</i>) DH5⍺ cells using the methods below.
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         <p> <i> Method 1: Converting between absorbance of cells to absorbance of a known concentration of beads </i>
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         <p> In the first method, silica beads are used to estimate the actual amount of cells during fluorescence measurement. These beads are modeled after a typical <i> E. coli </i> cell and are thus expected to scatter light in a similar way to <i> E. coli </i>          cells. As a sample of these silica beads gives a consistent and known absorbance measurement at 600 nm, absorbance measurements from a sample’s cell density can be converted into an “equivalent concentration of beads” measurement that should
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          be more universal and comparable between different labs.</p>
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        <p> <i>Method 2: Counting colony-forming units (CFUs) from the sample</i></p>
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        <p> In the second method, cell concentration is approximated is by plating a known volume of the sample and letting bacterial colonies grow. As each bacterial colony is assumed to represent a single cell (for cells that do not stick together), the
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          cell concentration in the sample is then directly proportional to the number of CFUs. Using a scaling factor computed from negative and positive control CFUs, a conversion factor from absorbance to CFU can be computed.</p>
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Revision as of 11:04, 15 October 2018

CONNECT WITH US

OVERVIEW

To write an overview.


To continue writing something else.




PARTS RECEIVED

Device Part Number Usage
Negative control BBa_R0040 TetR repressible promoter, medium strength promoter
Positive Control BBa_I20270 Promoter MeasKit (J23151)

EXPERIMENTS

Abs600
  • Wavelength: 600nm
  • Read Speed: Normal
  • Delay: 100 msec

Fluorescence
  • Excitation: 485
  • Emission: 525
  • Optics: Top
  • Gain: 50
  • Light Source: Xenon Flash
  • Lamp Energy: High
  • Read Speed: Normal
  • Delay: 100 msec
  • Read Height: 7 mm

You can put text here if you wanna

CONCLUSION


The results from our experiment seem to indicate that normalizing fluorescence measurements to absolute cell count using the Study’s two methods will not be able to reduce lab-to-lab variability because counting colony-forming units do not return the expected cell concentration, i.e. the cell concentration modeled by the silica beads in Method 1. While both methods cannot be used independently to establish a robust fluorescence measurement system, it may be possible that lab-to-lab variability can be reduced if a different method of normalizing to absolute cell count is devised, replacing Method 1, Method 2, or both.




HEAR OUR STORY

We’re using synthetic biology to change the way the world thinks and functions.

We’re a little disruptive, dangerous and maybe a tad too bold. But the world needs some shaking up every now and then, and we think we’re just the right people for that.

Together we’re going to engineer biology and create something awesome.

CONTACT US

nusigem@gmail.com

21 Lower Kent Ridge Rd, Singapore 119077