Team:Genspace/Collaborations
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We Helped Five other iGEM teams
1. UGA-Georgia iGEM Team
We helped the UGA-Georgia iGEM Team by characterizing their mCherry-based reporter system
The Genspace iGEM team helped the University of Georgia iGEM Team by participating in the Archaeal InterLab Study to further characterize the reproducibility of their mCherry reporter system. We received frozen cell lysates from them and followed the protocol below to develop the mCherry RFP overnight with shaking at 30C. The samples were then read in a plate reader with an excitation wavelength of 590nm and an emission wavelength of 645nm. The data were submitted to the UGA-Georgia team via their data submission form. A table of the data is presented below.
Here is the UGA Archaeal Collaboration Study Protocol:
As soon as you receive your samples in the mail, place them in a freezer (-80C) in order to reduce potential protein damages that may incur from freezing and thawing.
We recommend that you place your samples in the freezer and take time to read over our protocol as well as prepare your plan of action.
You have been given 30 total samples, (i.e., triplicate cell extracts obtained from 10 colonies, and please stay in accordance with our labels throughout your experiment.)
 
Each sample contains 10 ul of cell extracts. The buffer used is 25 mM PIPES pH 6.8. The PIPES should act as a negative control in your experiment. If PIPES is not available, a buffer with the same pH may be sufficient.
- Remove samples from the freezer and place in a shaker at 30C for overnight incubation. (This step is necessary because our cells are grown anaerobically but the fluorophore of mCherry fluorescent protein requires sufficient oxygen exposure for maturation.)
- Using “8B” determine the linear range for your measurement device.
- We recorded the highest fluorescence from sample “8B,” so if this is found to be in the linear range, then all the other samples will be as well.
- Our device required a 200 ul sample, so we found that a 100X dilution (i.e., 2 ul cell extracts re-suspended in 198 ul PIPES buffer) was within the linear range).
- Determining the linear range can be done with serial dilutions (e.g., 10, 10^2, 10^3, 10^4, and 10^5 dilutions).
- Our measurement device is a BioTek Synergy HT plate reader (96 well) and we use the Gen5 software.
- Excitation wavelength (nm): 590/20
- Emission wavelength (nm): 645/40
- Scaled* to High Wells
- Setpoint Temperature: 25C
- Your measurement device and measurement protocol may vary from ours and other teams, so please update our form with your specific parameters. Email Steven Kodish at skodish@uga.edu, Rebecca Ann Buchanan beccab@uga.edu, and Akshay Chandora at akshaych@uga.edu upon receiving this shipment for the link to the registration form and data form or for any additional questions.
*Our values are all relative to each other (i.e., X is 20% more fluorescent than Y) so that all data can be correlated regardless of measurement device.
2. University of Michigan iGEM Team
We helped the University of Michigan team in their effort to build an online protocol database called ProtoCat as our project for the iGEM competition. To help them better understand what should go in to ProtoCat, we participated in a survey where we answered questions about how we find and use protocols.
3. Paris-Bettencourt iGEM Team
We helped the Paris-Bettencourt team by adding our project data to their Rhizi Database.
4. Columbia University iGEM Team
We helped the Columbia University team by lending them critical reagents.
5. Cooper Union iGEM Team
We helped the Cooper Union team by lending them critical reagents and equipment.