Interlab Measurement Study

The Interlab Study is an initiative started in 2014 by the iGEM foundation to collect standard measurement data from around the world. This year, as a part of the measurement track, we completed the Interlab study according to the directions provided on the iGEM Interlab Study webpage. The purpose of this year's study was to measure the absorbance and fluorescence of three different GFP containing devices as well as a positive and negative control. The absorbance and fluorescence of a series of diluted FITC samples and LUDOX-S30 to serve as measurement standards. Below, we have detailed our procedures, implications of our results, and conclusions we have drawn that relate back to our main focus of evolutionary stability in synthetic biology. In synthetic biology, it is important that part characterization is consistent between different labs to create well-defined standard parts for the Registry. We are excited to contribute to this study!

Protocol

1. Transform the genetic parts from the Interlab 2016 kit into TOP10 E. coli cells
2. Streak out an agar plate (LB/CAM) with the E. coli containing the device and any controls
   • Streak out one plate/device and control
   • Incubate plates overnight (18-20 hours) at 37°C
3. Inoculate liquid culture with experimental devices (in duplicate) and controls in test tube in 10 ml LB/CAM media (2 test tubes for each device for a total of 10)

Plate Reader Protocol

1. Obtain an initial OD600 measurement of the overnight cultures, based on this data dilute the samples to a target OD600 measurement of around 0.02
2. Incubate liquid cultures for 6 hours at 300 rpm in 37°C. At each hour time point, starting with 0 hour, a 100 µl sample of each culture was loaded onto a clear-bottom 96 well plate, this ended after 6 hours
3. The samples were measured using a Infinite 200 Pro plate reader. The settings were: Excitation at 495 nm with a 9 nm bandwidth and Emission at 530 nm with a 20 nm bandwidth. The OD600 was also measured

Ludox Measurement