Interlab Measurement

As a team under measurement track, we are delighted to participate in the Interlab Measurement Study. The objective of this study is to measure fluorescence data for three GFP-expressing plasmid devices and by collecting all the data gathered from the participated iGEM teams all over the world, we are able to assess the precision and reproducibility of the results from different labs.

The three constructs are differentiated by their promoters and all the rest of the parts are the same. Hence, we would be able to determine the differences in terms of the strength of these promoters. The three promoters, namely J23101, J23106, J23117, were cloned into pSB1C3 vector and submitted by iGEM Team Berkley in 2006. Compared to the Interlab Study performed in the previous years, two controls both positive (I20270) and negative (R0040) have also been added into the measurement test, used as reference devices.

Using water absorbance as a blank, the corrected absorbance of LUDOX is calculated as 0.00415. Correction factor was then calculated by dividing the corrected absorbance by a reference OD value 0.01475. Hence, the correction factor is 3.554.

OD chart

Figure 3. Absorbance measurement of LUDOX 100% and H2O and correction factor table

FI represents the fluorescence intensity and absorbance 600 represents the density of bacterial culture. Therefore, FI/Abs600 is used to assess the strength of the GFP expression.


Figure 4. FITC standard curve plotted from the data of 12 dilutions of concentration of 4 replicates


Figure 5. Flouresence intensity plot (FI) of 5 devices and each with 2 replicates from 0-6h. From the plot, we could see that the GFP expression of the first two devices are much higher than the rest.


Figure 6. Absorbance at 600nm of the 5 devices from two replicates. All the cell culture have similar growth trend except for device 1 replicate. We speculate that it might be due to the high level of GFP expression slows down the growth.


Figure 7. FI/Abs ratio of the replicates and devices from 0-6h. Device 1 has the highest level in terms of Floresence expression and Device 3 has the lowest among the 3 devices being tested. (Note: The error bar is different from the Excel file we have submitted because there is an error in the Excel template that has been provided in the website. And this plot is the corrected one.)

From the plots above, we could see that Device 1 has the strongest FI and highest GFP expression, followed by device 2 which has similar data close to the positive control. Furthermore, Device 3 has the lowest GFP expression and therefore it has the weakest promoter. In summary, we conclude that the promoter strength of the three constructs: Device 1> Device 2> Device 3.

Beal, J., Haddock-Angelli, T., Gershater, M., de Mora, K., Lizarazo, M., Hollenhorst, J., & Rettberg, R. (2016). Reproducibility of Fluorescent Expression from Engineered Biological Constructs in E. coli. PloS one, 11(3), e0150182.