Team:NWU-CHINA/InterLab
Interlab Study
Overview
As GFP is an useful indicator, we are delighted to participate in the Interlab Measurement Study to help HQ researching the differences of results from all participated IGEM team all over the world.
This year, we measured 6 devices and two controls. There are totally 3/5 promoters, which were combined differently in different device. Therefore, we can measure different result caused by different combination. The three promoters, namely J23101, J23106, J23117, were cloned into pSB1C3 vector and submitted by Igem Team Berkley in 2006. The two controls, I20270 and R0040, were used as reference device.
Experiment
Firstly, We test our competent cells as HQ requirement. We followed protocol on the competent cells test page. Their competent met requirement of competent cells.
Detail data is shown as follows:
Then we transferred devices into our competent cells. At the beginning, because our chlor lost efficiency, our cell couldn’t be selected.
After changing antibiotic, we conducted our Interlab Study fomally.
Results
We did this experiment twice. For the first time, we didn’t get ideal data. Our cells’ fluorescence reading didn’t increase with time passing. It means that our cell didn’t express the device.
For the second time, we got data relevant with time. However, our reading value is really low. We used agarose gel electrophoresis to detect our gene expression condition.
As the picture shows, every device have transferred into chassis successfully. Therefore, we think that the reason our reading value low is because of our measuring setting. Our gain setting is 35, which leads to a low reading value. We also agree that this setting influence results a lot. We found that blank well of plate also have reading. Our low reading setting may let these blank well reading influence our result.
Our first experiment result is not good enough, so we won’t show it off. Our second experiment results are as follows:
Although the fluorescence reading is low, it still shows a obvious relevance with time.
During our experiment, we found that device 1, device 3, device 6 didn’t express well. However, when putting cells under UV, Device 1 shows obvious fluorescence, while device 3 and device 6 still have no phenomenon.
Our Abs reading and cell culture results are as follows:
When we contrasted the regulator and RBS upstream of GFP. B0034 and J3641100 worked in other devise, so we came to a conclusion that J23117 or the combination of J23117 and B0034 or J364100 may has some negative effect to the expression of device, while the combination of J23101 and B0034 can cause a slow growth rate to cell.
As our gain setting value is 35, our fluorescence reading is low. we think that data measured in low gain setting are not accurate enough. We think our data can be a good sample for other teams.
