Difference between revisions of "Part:BBa K1493501:Experience"

 
(→‎User Reviews)
Line 19: Line 19:
 
<!-- End of the user review template -->
 
<!-- End of the user review template -->
 
<!-- DON'T DELETE --><partinfo>BBa_K1493501 EndReviews</partinfo>
 
<!-- DON'T DELETE --><partinfo>BBa_K1493501 EndReviews</partinfo>
 +
 +
 +
[http://2015.igem.org/Team:Harvard_BioDesign Harvard BioDesign 2015] used the rhamnose promoter to express fimH adhesin in [https://parts.igem.org/wiki/index.php?title=Part:BBa_K1850000 BBa_K1850000] and BBa_K1850002-11. We were interested in maintaining very tight control over the expression of this protein to allow for control over the strength of binding and avoid gene-dosing issues. We selected the rhamnose promoter for this purpose, but were interested in a hypothesized characteristic of the promoter, which is that it is titratable on a colony level, but not on the level of a single cell. In other words, each single cell is either completely "on" or completely "off", but on a population level this ratio changes and appears titratable on the average. To that end we performed Fluorescence-activated cell sorting with using a flow cytometer, with variable inducer quantities, to characterize fluorescence for single cells. We found that the rhamnose promoter can either activate florescence completely or not at all, as shown in the bimodal fluorescence distribution of the samples:
 +
 +
[https://static.igem.org/mediawiki/2015/4/4c/Harvard_GFP_Rham.png]
 +
 +
This confirms the hypothesis that the rhamnose promoter is not titratable on a single cell level. We believe this information useful in understanding induction results for systems which depend on on close control of protein expression.

Revision as of 04:43, 25 September 2015


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_K1493501

User Reviews

UNIQdc46ac5367953ffd-partinfo-00000000-QINU UNIQdc46ac5367953ffd-partinfo-00000001-QINU


[http://2015.igem.org/Team:Harvard_BioDesign Harvard BioDesign 2015] used the rhamnose promoter to express fimH adhesin in BBa_K1850000 and BBa_K1850002-11. We were interested in maintaining very tight control over the expression of this protein to allow for control over the strength of binding and avoid gene-dosing issues. We selected the rhamnose promoter for this purpose, but were interested in a hypothesized characteristic of the promoter, which is that it is titratable on a colony level, but not on the level of a single cell. In other words, each single cell is either completely "on" or completely "off", but on a population level this ratio changes and appears titratable on the average. To that end we performed Fluorescence-activated cell sorting with using a flow cytometer, with variable inducer quantities, to characterize fluorescence for single cells. We found that the rhamnose promoter can either activate florescence completely or not at all, as shown in the bimodal fluorescence distribution of the samples:

[1]

This confirms the hypothesis that the rhamnose promoter is not titratable on a single cell level. We believe this information useful in understanding induction results for systems which depend on on close control of protein expression.