Composite

Part:BBa_K410000:Experience

Designed by: Christina Graves, Scott Holmes   Group: iGEM10_GeorgiaTech   (2010-10-27)
Revision as of 08:43, 29 September 2011 by Egaruz89 (Talk | contribs) (→‎Methodology)


This experience page is provided so that any user may enter their experience using this part. Please enter how you used this part and how it worked out.

User Reviews

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egaruz89

We expected the part to have a better development at lower temperatures than 37°C, like at 23°C, but the result is almost the same as in other parts not designed for that. Our overall review is 3/5

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Experiment Objectives

1. Test the BioBrick operation to different substrates and temperatures, using KNO3 40 mM: a.Substrates:

  • LB
  • Minimal essential medium
  • Saline solution
  • Lunar Land

b.Temperatures:

  • 8°C
  • 23°C
  • 37°C

2. Improve the characterization of the BioBrick tested in other substrates to study the performance of the promoter.
3. Define the Biobrick promoter strength at the temperatures and selected substrates.

Methodology

We chose 4 temperatures:

  • 5-10°C
  • 20-30°C
  • 37°C

These temperatures were selected so we could evaluate the response to cold shock in various ranges of our BioBrick. Since the lab equipments we had available couldn’t reach temperatures between 11-20°C this temperature was not used during the experience. Finally, the lab equipment allowed us to make the experiments in: 37°C, 23°C and 8°C. We also chose 3 different Growth Media:

  • Saline Solution
  • Liquid LB
  • Minimal Media

These growth medias where selected to evaluate the response of our BioBricks in different metabolic pathways. According to the characterization made by Team Edinburgh 2009, the Pyear promoter (BBa_K216005) used by Bristol 2010, works best in minimal media and complex media with a nitrate concentration of 40mM. Using this reference, we added 40mM of KNO3 to every growth media in the different temperatures for Bristol 2010 - BBa_K381001 and UTP-Panama 2011 – BBa_K672000.
We prepared 9 15mL Falcon Tubes, these tubes contained:

  • First 3 (one for each temperature): 10mL of Saline Solution + a colony of the BioBrick + 40mM of KNO3.
  • Second 3 (one for each temperature): 10mL of Minimal Media + a colony of the BioBrick BioBrick + 40mM of KNO3.
  • Lat 3 (one for each temperature): 10mL of Liquid Luria Broth + a colony of the BioBrick BioBrick + 40mM of KNO3.

Results

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