Part:BBa_K2598043
amilCP+amilGFP
This part contains two chromoproteins amilCP and amilGFP. We use thiss part to mix colors and get a more diverse Chromatography。 It is part of the GRB system, see more information from RGB System
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Characterization
Figure 1 shows the relationship between the wavelength of light exposed on liquid medium and the intensity of BFP, GFP and RFP E. coli expressed from left figure to right figure respectively. We got the data through flow cytometer and analyzed it to get the figure. The y-axis is the number of cells, and the x-axis is fluorescence intensity. And every color is E. coli that grows for 8 hours under the light of the corresponding wavelength. We can see E. coli has the highest blue fluorescence expression under blue light from the left graph. And We can also see E. coli has the highest green and red fluorescence expression under green light and right light from the middle and right graph respectively. So this figure proves that our system and our parts can work well.
Figure 2 shows the relationship between fluorescence intensity and excitation wavelength. The x-axis is wavelength of 10h illumination. The solid medium gradually emerged and the y-axis is RGB figure of fluorescence in illuminated solid medium. This curve illustrates how our system responses to different excitation wavelength, which perfectly meets our expectation. So this figure proves that our system and our parts can work well.
Figure 3 shows colors we got from the solid medium exposed under light, in which E. coli producing fluorescent protein grows. When E .coli producing fluorescent protein are exposed under uniform light of single wavelength, the solid medium gradually emerged corresponding colors. And using color picker, we get many pure colors with predominant continuity.
We explored the relationship between fluorescent intensity and illumination intensity, which affects the shade of the color.
Figure 4 shows the red fluorescent intensity of E. coli under light of 620-630nm wavelength with different illumination intensity. We found when illuminated under around 961lux light, we can get the most red fluorescence. We also explore the relationship between green and blue fluorescent intensity of E. coli under light of 515-530nm wavelength and 460-470nm wavelength respectively and illumination intensity. The results are similar, that is, moderate intensity of light is most favorable for E. coli to express fluorescence.
Figure 5 shows results of gel electrophoresis of various parts after PCR with primer VF2\VR. The distance between primer binding sites and both ends of the parts are approximately 150 bp, thus rendering the product about 300 bp longer. The picture is edited to show a more compact photo.
Figure 6 shows the tandem expression of chromoproteins. By putting two chromoprotein RBSs and genes under one promoter, we constructed six plasmids(BBa_K2598043, BBa_K2598044, BBa_K2598045, BBa_K2598046, BBa_K2598047, BBa_K2598048) to tandem express eforred, amilCP, amilGFP and fwYellow chromoproteins. As shown in the figure, we built a color spectrum using chromoproteins and their tandem expression products, which can show orange, pink, yellow and green colors. Furthermore, we can see that the color of the tandem expression products, is always between the colors of the two chromoproteins in the spectrum. That is to say, by applying the physical principles, we can easily mix the color we want using tandem expression of chromoproteins.
Figure 7 shows the tandem expression of two chromoproteins， amilCP and amilGFP, which are blue and yellow respectively. We mixed blue and yellow together to create green color, which rarely exist in colors of chromoproteins in iGEM registry. We use color picker and Photoshop to analyze and compare the color we got and that of amilGFP. The RGB value of our color is 184-209-108, while that of amilGFP is 178-191-138. That shows our color from the tandem expression is brighter and greener.
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