Difference between revisions of "Team:NAWI Graz/Results"
| Line 1: | Line 1: | ||
{{NAWI_Graz:navbar}} | {{NAWI_Graz:navbar}} | ||
<html> | <html> | ||
| − | |||
<div class="jumbotron"> | <div class="jumbotron"> | ||
<div class="section section-heading container"> | <div class="section section-heading container"> | ||
| Line 7: | Line 6: | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | <div class="section container"> | |
| − | + | <h2 class="section-sub">pH-Sensing</h2> | |
| − | + | <div class="section-text container"> | |
| − | <p>To test if our pH sensitive constructs express the fluorescence proteins we cultivated the bacteria hosting the plasmids in LPM with pH 7 overnight. For expression control for our alx-mNeonGreen construct, we inoculated 20 ml LPM with pH 7.0, 8.0 and 8.5 to an OD<sub>600</sub> of 0.2 using the overnight culture. After 20 and 40 minutes 1 ml of each culture where taken and adjusted to the lowest OD<sub>600</sub> of the three samples to standardize OD<sub>600</sub>. This was necessary growth at pH 8.0 and 8.5 was significant slower than at pH 7.0. This diluted samples where used to measure fluorescence (extinction 490 nm, absorbance 520 nm) and again OD<sub>600</sub> with our plate reader. Three aliquots of each sample where measured (n=3), fluorescence was divided by OD<sub>600</sub> and average of the three aliquots where taken to obtain the data shown in Figure 3.</p> | + | <p>To test if our pH sensitive constructs express the fluorescence proteins we cultivated the bacteria hosting the plasmids |
| − | + | in LPM with pH 7 overnight. For expression control for our alx-mNeonGreen construct, we inoculated 20 ml LPM | |
| − | + | with pH 7.0, 8.0 and 8.5 to an OD | |
| − | + | <sub>600</sub> of 0.2 using the overnight culture. After 20 and 40 minutes 1 ml of each culture where taken and adjusted | |
| − | + | to the lowest OD | |
| − | + | <sub>600</sub> of the three samples to standardize OD | |
| − | + | <sub>600</sub>. This was necessary growth at pH 8.0 and 8.5 was significant slower than at pH 7.0. This diluted samples | |
| − | + | where used to measure fluorescence (extinction 490 nm, absorbance 520 nm) and again OD | |
| − | + | <sub>600</sub> with our plate reader. Three aliquots of each sample where measured (n=3), fluorescence was divided | |
| − | + | by OD | |
| + | <sub>600</sub> and average of the three aliquots where taken to obtain the data shown in Figure 3.</p> | ||
| + | </div> | ||
| + | <div class="row container"> | ||
| + | <div class="col"> | ||
| + | <img class="section-image" src="" alt="[Diagramm_alx.xlsx]"> | ||
</div> | </div> | ||
| − | + | <div class="col image-subtext border border-secondary"> | |
| − | + | <font size="-2"> | |
| − | + | <b>Fig 1. Alx controlled mNeonGreen Expression: </b>Culture media was bufferd to pH 7, 8 and 8.5 and inoculated | |
| − | < | + | to an OD |
| + | <sub>600</sub> of 0.2. After 20 and 40 minutes, aliquotes where taken and fluorescence (extinction 490 nm, absorbance | ||
| + | 520 nm) and OD | ||
| + | <sub>600</sub> were measured with the plate reader (n=3).</font> | ||
| + | <br> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="section-text container"> | |
| − | + | As shown in figure 1. Expression of mNeonGreen is increased at pH 8 and 8.5 compared to standard pH 7. Cultures | |
| − | + | at ph 7, but also cultures without the mNeonGreen plasmid (data not shown) show a high basic fluorescence, leading | |
| − | + | to the conclusion that the cells naturally show emission at 520 nm. Still, fluorescence is increased twofold at pH | |
| − | < | + | 8.5 after 20 and 40 minutes and fluorescence is significantly increased at pH 8 at both measuring points |
| − | < | + | </div> |
| − | As shown in figure 1. Expression of mNeonGreen is increased at pH 8 and 8.5 compared to standard pH 7. Cultures at ph 7, but also cultures without the mNeonGreen plasmid (data not shown) show a high basic fluorescence, leading to the conclusion that the cells naturally show emission at 520 nm. Still, fluorescence is increased twofold at pH 8.5 after 20 and 40 minutes and fluorescence is significantly increased at pH 8 at both measuring points | + | </div> |
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
</html> | </html> | ||
{{NAWI_Graz:footer}} | {{NAWI_Graz:footer}} | ||
Revision as of 21:47, 31 October 2017
RESULTS
pH-Sensing
To test if our pH sensitive constructs express the fluorescence proteins we cultivated the bacteria hosting the plasmids in LPM with pH 7 overnight. For expression control for our alx-mNeonGreen construct, we inoculated 20 ml LPM with pH 7.0, 8.0 and 8.5 to an OD 600 of 0.2 using the overnight culture. After 20 and 40 minutes 1 ml of each culture where taken and adjusted to the lowest OD 600 of the three samples to standardize OD 600. This was necessary growth at pH 8.0 and 8.5 was significant slower than at pH 7.0. This diluted samples where used to measure fluorescence (extinction 490 nm, absorbance 520 nm) and again OD 600 with our plate reader. Three aliquots of each sample where measured (n=3), fluorescence was divided by OD 600 and average of the three aliquots where taken to obtain the data shown in Figure 3.
Fig 1. Alx controlled mNeonGreen Expression: Culture media was bufferd to pH 7, 8 and 8.5 and inoculated
to an OD
600 of 0.2. After 20 and 40 minutes, aliquotes where taken and fluorescence (extinction 490 nm, absorbance
520 nm) and OD
600 were measured with the plate reader (n=3).
As shown in figure 1. Expression of mNeonGreen is increased at pH 8 and 8.5 compared to standard pH 7. Cultures
at ph 7, but also cultures without the mNeonGreen plasmid (data not shown) show a high basic fluorescence, leading
to the conclusion that the cells naturally show emission at 520 nm. Still, fluorescence is increased twofold at pH
8.5 after 20 and 40 minutes and fluorescence is significantly increased at pH 8 at both measuring points
