Part:BBa_K412000:Experience

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Applications of BBa_K412000

• All images were taken with Olympus IX81 automated inverted microscope specially equipped for live cell imaging. All experiments were performed with DH5α E.coli cells. The filter sets we used are: 545/30x (excitation) and 620/60m (emission) filters for DsRed, 470/40x (excitation) and 525/50m (emission) for GFP. Data collection and processing was performed by the SlideBook software.
• The data was taken using cells that were grown overnight in 2mL of LB broth that had 50ug/mL of the appropriate antibiotic at 37C and 220rpm. This was used in a 1:50 dilution. Each sample had 40uL from the broth grown overnight and 2mL of the LB broth with the appropriate antibiotic. These were allowed to grow until a predetermined OD. Then the contaminants were added and the cells were allowed to grow for another 2 hours, before the images were taken.
• The images shown are the images used to get the data. From left to right the image is the negative control to a 10mM concentration of the respective contaminants. The graph shows the average of all the maximum intensities of the cells in the image vs the concentration of nitrate/nitrite added to the sample. The data shows that PyeaR+GFP responds better to nitrates than nitrites. It also shows that the cell is more sensitive to aluminum nitrate nonahydrate, and therefore starts dying at a lower concentration.

The images above are timelapse images. The images were taken every ten minutes after the 10mM potassium nitrate was added to the sample.

User Reviews

UNIQ1b5289837dcb29ec-partinfo-00000000-QINU UNIQ1b5289837dcb29ec-partinfo-00000001-QINU

••••• iGEM12_OUC-China

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Results of nitrate induction are presented on the left. (A) It is obvious that nitrate can induce GFP expression. Additionally, Pyear (nitrate-sensitive promoter) respond ultra-sensitively when nitrate concentration ranges from approximately 0-2 ug/L. And after concentration of 3.5ug/L,GFP expression maintains at nearly 2800 RFU which showcase its maximal initiation rate of promoter. (B)Top10 brand control is presented. We can clearly see the differences between (A) and (B).