Difference between revisions of "Part:BBa K895007:Experience"

(→‎User Reviews)
(→‎Results)
Line 22: Line 22:
  
 
===Results===
 
===Results===
−
<i>E. coli</i> MG1655 was transformed with <partinfo>BBa_K895007</partinfo> grown aerobically overnight in LB medium. The culture was then used to inoculate 30 ml anaerobic cultures in LB only, LB + 0.4% (w/v) tetrathionate or LB + 0.4% (w/v) thiosulfate. Following 16 hours growth aliquotes were taken and analysed by SDS-PAGE and Western immunoblotting using an anti-GFP monoclonal.
+
<b><u>Characterisation by Dundee iGEM Team 2012</u></b>
 +
<p> Tetrathionate reductase is an enzyme found in most <i>Salmonella</i> strains and some other opportunistic pathogens such as <i>Serrata</i>. The expression of the tetrathionate reductase genes (<i>ttrBCA</i>) in <i>Salmonella</i> is regulated by a two-component system. This two-component system senses extracellular tetrathionate <i>via</i> a membrane-bound histidine kinase (TtrS), which in turn serves to promote phosphorylation of the DNA-binding TttR response regulator (Hensel <i>et al</i>. 1999). It is possible that this two-component system could be used to regulate any reporter protein or other enzyme system.</p>
 +
<p>
 +
<p><i>E. coli</i> MG1655 was transformed with <partinfo>BBa_K895007</partinfo> grown aerobically overnight in LB medium. The culture was then used to inoculate 30 ml anaerobic cultures in LB only, LB + 0.4% (w/v) tetrathionate or LB + 0.4% (w/v) thiosulfate. Following 16 hours growth aliquots were taken and analysed by SDS-PAGE and Western immunoblotting using an anti-GFP monoclonal.</p>
  
−
This small scale aassay reveals that GFP was only expressed from the plasmid in the presence of tetrathionate. The device therefore works as a putative <b>inflammation biosensor</b>.
+
<p>This small scale aassay reveals that GFP was only expressed from the plasmid in the presence of tetrathionate. The device therefore works as a putative <b>inflammation biosensor</b>.</p>
  
 
[[Image:Dundee_Biosensor_Figure5.jpg]]
 
[[Image:Dundee_Biosensor_Figure5.jpg]]

Revision as of 09:03, 29 September 2012

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_K895007

User Reviews

UNIQ06d8b9db5bc3f238-partinfo-00000000-QINU

•••••

iGEM Dundee 2012

This part was seen work in practice. The device produces GFP in response to extracellular tetrathionate (Figure 1). The device is not activated by thiosulfate, which is the natural product of tetrathionate reduction (Figure 1).

UNIQ06d8b9db5bc3f238-partinfo-00000002-QINU

Results

Characterisation by Dundee iGEM Team 2012

Tetrathionate reductase is an enzyme found in most Salmonella strains and some other opportunistic pathogens such as Serrata. The expression of the tetrathionate reductase genes (ttrBCA) in Salmonella is regulated by a two-component system. This two-component system senses extracellular tetrathionate via a membrane-bound histidine kinase (TtrS), which in turn serves to promote phosphorylation of the DNA-binding TttR response regulator (Hensel et al. 1999). It is possible that this two-component system could be used to regulate any reporter protein or other enzyme system.

<p>E. coli MG1655 was transformed with BBa_K895007 grown aerobically overnight in LB medium. The culture was then used to inoculate 30 ml anaerobic cultures in LB only, LB + 0.4% (w/v) tetrathionate or LB + 0.4% (w/v) thiosulfate. Following 16 hours growth aliquots were taken and analysed by SDS-PAGE and Western immunoblotting using an anti-GFP monoclonal.

This small scale aassay reveals that GFP was only expressed from the plasmid in the presence of tetrathionate. The device therefore works as a putative inflammation biosensor.

Dundee Biosensor Figure5.jpg

Figure 1: Production of GFP from BBa_K895007 is tetrathionate dependent. The RED ARROW points to the lane producing GFP.



References

UNIQ06d8b9db5bc3f238-partinfo-00000005-QINU