Team:Nankai/Measurement
Is measurement important?
Precise measurement is crucial since it is the foundation of every scientific discipline. More importantly, measurement can also be an act of communication if various laboratories around the world collaborate together and make meaningful comparisons between same or similar observations.
How can we help?
The goal of this year’s interlab study tract is to compare the ability of transcription initiation of three different promoters called P1, P2 and P3. These three promoters are constitutive promoters of E. coli and their ability of transcription initiation has already been determined by RFP reporter gene. We're going to measure the ability of each promoter by another way.
What did we do?
In our experiment, we use the coding gene of Green Fluorescence Protein as a new reporter gene. First we successfully constructed three recombinant plasmids, in which the three promoters are located upstream of the GFP coding gene. Then these recombinant plasmids were transformed into the E. coli cells and several strains that steadily express GFP were obtained.
After the transformed strains were obtained, they were firstly observed under a fluorescent microscopy.Next, to confirm our result quantitatively, we used a flow cytometry to measure the average fluorescence intensity of each strain, as instructed.We also made a figure of the average fluorescence intensity to show the different transcription initiation ability of each promoter more clearly.
And the result?
The results are as following:
Fig.1 Fluorescent Microscopy Picture of Transformed E. coli
Fig.2 Fluorescence Intensity of Transformed E. coli
Fig.3 Average Fluorescence Intensity of Transformed E. coli
Our conclusion
Fig.3 Average Fluorescence Intensity of Transformed E. coli
Our conclusion
From the results we can clearly see that the ability of P1 is the strongest while the ability of P3 is the weakest. The ability of P2 is in the middle. Also, the differences of these three promoters are statistically significant.So our final conclusion is that our measurement result is basically in consistent with the official data, i.e. the data from the RFP reporter gene.
.
Flow cytometry was used to measure the average fluorescence intensity of each strain.
Centrifuge was used to seperate matarials and precipitate E. coli.
In the progress of transferring E. coli suspension.
In the progress of extracting plasmids.
Result of the concentration of plasmids.