Difference between revisions of "Part:BBa K302012:Experience"
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We characterised the part first without, and then with, LacI repression (using the integration vector pMutin4 to integrate ''lacI'' into the ''Bacillus subtilis'' 168 chromosome). | We characterised the part first without, and then with, LacI repression (using the integration vector pMutin4 to integrate ''lacI'' into the ''Bacillus subtilis'' 168 chromosome). | ||
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|with inhibition cell lengths are comparable to ''Bacillus subtilis 168'' at 0μM IPTG and longer with IPTG induction. | |with inhibition cell lengths are comparable to ''Bacillus subtilis 168'' at 0μM IPTG and longer with IPTG induction. | ||
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Revision as of 17:47, 27 October 2010
This experience page is provided so that any user may enter their experience using this part.
Please enter
how you used this part and how it worked out.
Applications of BBa_K302012
Characterisation by Team Newcastle 2010
We integrated our part into the Bacillus subtilis 168 chromosome at amyE (using the integration vector pGFP-rrnB) and selected for integration by testing for the ability to hydrolyse starch. Homologous recombination at amyE destroys endogenous expression of amylase. Colonies that are not able to break down starch on agar plate do not have a white halo when exposed to iodine.
The part was co-transcribed with gfp fluorescent marker by transcriptional fusion after the yneA coding sequence.
We characterised the part first without, and then with, LacI repression (using the integration vector pMutin4 to integrate lacI into the Bacillus subtilis 168 chromosome).
Table1:
| Stats: | 168 | yneA | pMutin4 0μM IPTG | pMutin4 1μM IPTG |
|---|---|---|---|---|
| Average: | 1.34μm | 3.53μm | 1.74μm | 3.19μm |
| Max: | 2.30μm | 6.00μm | 3.62μm | 9.77μm |
| Min: | 0.55μm | 1.31μm | 0.88μm | 1.14μm |
| Median: | 1.33μm | 3.27μm | 1.62μm | 2.66μm |
| Standard Deviation: | 0.32μm | 1.01μm | 0.80μm | 1.56μm |
Figure1:
| Distribution of cell lengths is not normal, so the mean is misleading; we are reporting the median instead. |
|
Figure1: shows statistics for populations of cells
|
| with inhibition cell lengths are comparable to Bacillus subtilis 168 at 0μM IPTG and longer with IPTG induction. |
Figure2:
  
|
| Figure2: Bacillus subtilis 168 cells (left),Bacillus subtilis expressing yneA(centre) and Bacillus subtilis overexpressing yneA(right) |
| The images we have taken this data from had very different numbers of cells, so the cells counts are misleading therefore we are reporting the proportions of cells at a given length. |
Figure 3:
|
| Figure 3 shows the percentage of cells at different lengths (μm) uninduced |
Figure 4:
| Figure 4:Bacillus subtilis 168 cells (left) and non-induced cells (right) |
|
Figure 5:
|
| Figure 5 shows the percentage of cells at different lengths(μm)induced at 0.2mM IPTG |
Figure 6:
|
| Figure 6: Bacillus subtilis 168 cells (left) and cells induced at 0.2mM IPTG (right) |
Figure 7:
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| Figure 7: shows the percentage of cells at different lengths (μm) induced at 1mM IPTG |
Figure 8:
|
| Figure 8: Bacillus subtilis 168 cells (left) and cells induced at 1mM IPTG(right) |
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