Difference between revisions of "Team:Jilin China/InterLab"

Reliable and repeatable measurement is a key component to all engineering disciplines. The same holds true for synthetic biology, which has also been called engineering biology. However, the ability to repeat measurements in different labs has been difficult. The Measurement Committee, through the InterLab study, has been developing a robust measurement procedure for green fluorescent protein (GFP) over the last three years. We chose GFP as the measurement marker for this study since it's one of the most used markers in synthetic biology and, as a result, most laboratories are equipped to measure this protein. We think it is a great chance for our team to make our own contribution to this project.

• Positive Control (BBa_I20270): well 20B
• Negative Control (BBa_R0040): well 20D
• Test Device 1 (BBa_J364000): well 20F
• Test Device 2 (BBa_J364001): well 20H
• Test Device 3 (BBa_J364002): well 20J
• Test Device 4 (BBa_J364003): well 20L
• Test Device 5 (BBa_J364004): well 20N
• Test Device 6 (BBa_J364005): well 20P

We strictly followed the protocol provided by iGEM. We measured the experiment of OD600 reference point and fitted the fluorescence standard curve first and then detected GFP expression in our E.coli with the plate reader Synergy HT from BioTek.

We have sent our data sheets to measurement AT igem DOT org before 29th September.

The sheets are as followed:

1.OD600 reference point


2.Fluorescein standard curve




3.Raw Plate Reader Measurements
Dilution Calculation:


Fluorescence Raw Readings:

Abs600 Raw Readings

fig.1a


fig.1b

fig.1a&b The absorbance at 600nm over time. Samples were collected every two hours and measured Abs600 by using the plate reader.

fig.2a


fig.2b

fig.2a&b The change of fluorescence over time. Samples were collected every two hours and measured their fluorescence by using the plate reader.

fig.3 The value of Fluorescein/OD600 of every test device in different time.

As our results showed, promoter J23101 was the strongest among the 3 promoters, which showed the highest GFP fluorescence. Promoter J23106 was moderate, and promoter J23117 was the lowest.

We really appreciated Prof. Yubin Ge for providing us the permission to use the plate reader from his lab. Besides, PhD candidate Cheng Hu taught us how to use a plate reader. BioTek service engineer Jeremy Gagne told us some settings of the Synergy HT, which we couldn't find from the setting panel.