Difference between revisions of "Part:BBa K2992001"
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===Usage and Biology=== | ===Usage and Biology=== | ||
− | P<i>ntnh</i> is found upstream of the 5'-UTR and RBS of the non-toxic non-haemagglutinin component (ntnh) of the neurotoxin gene cluster in <i>C. botulinum</i>. This promoter was used to drive expression of | + | P<i>ntnh</i> is found upstream of the 5'-UTR and RBS of the non-toxic non-haemagglutinin component (ntnh) of the neurotoxin gene cluster in <i>C. botulinum</i>. This promoter was used to drive expression of our chosen reporter genes of interest. Doing so, allowed us to link BotR expression with the transcriptional activation of our reporter genes. |
===Characterisation=== | ===Characterisation=== | ||
− | This basic part was used for the assembly of our composite parts and characterised | + | This basic part was used for the assembly of our composite parts and characterised using 3 difference reporters: [https://parts.igem.org/Part:BBa_K2992039 GusA (BBa_K2992039)], [https://parts.igem.org/Part:BBa_K2992044 FAST (BBa_K2992044)] and [https://parts.igem.org/Part:BBa_K2992036 acetone (BBa_K2992036 )]. More information can be found on our [https://2019.igem.org/Team:Nottingham/Results Results Page].<br> <br> <br> |
<br> | <br> | ||
https://2019.igem.org/wiki/images/5/5e/T--Nottingham--Basic4.png | https://2019.igem.org/wiki/images/5/5e/T--Nottingham--Basic4.png | ||
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https://2019.igem.org/wiki/images/6/6b/T--Nottingham--Basic3.png | https://2019.igem.org/wiki/images/6/6b/T--Nottingham--Basic3.png | ||
<br> | <br> | ||
− | Characterisation of this promoter against P<i>fdx</i>, P<i>thl</i> and P<i>botR</i> using FAST fluorescent assay, showed P<i>ntnh</i> | + | Characterisation of this promoter (comprising parts [https://parts.igem.org/Part:BBa_K2992001 BBa_K2992001] and [https://parts.igem.org/Part:BBa_K2992015 BBa_K2992015]) against P<i>fdx</i>, P<i>thl</i> and P<i>botR</i> using FAST fluorescent assay, showed that P<i>ntnh</i> is a medium-strength promoter in E.<i>coli</i>, while being very weak in C.<i>sporogenes</i> (only slightly stronger than the promoterless control), as expected in the absence of BotR. <br> |
Latest revision as of 02:55, 22 October 2019
PntnH
Promoter region for the non-toxic non-haemagglutinin component of the neurotoxin gene cluster in Clostridum botulinum.
Usage and Biology
Pntnh is found upstream of the 5'-UTR and RBS of the non-toxic non-haemagglutinin component (ntnh) of the neurotoxin gene cluster in C. botulinum. This promoter was used to drive expression of our chosen reporter genes of interest. Doing so, allowed us to link BotR expression with the transcriptional activation of our reporter genes.
Characterisation
This basic part was used for the assembly of our composite parts and characterised using 3 difference reporters: GusA (BBa_K2992039), FAST (BBa_K2992044) and acetone (BBa_K2992036 ). More information can be found on our Results Page.
Characterisation of this promoter (comprising parts BBa_K2992001 and BBa_K2992015) against Pfdx, Pthl and PbotR using FAST fluorescent assay, showed that Pntnh is a medium-strength promoter in E.coli, while being very weak in C.sporogenes (only slightly stronger than the promoterless control), as expected in the absence of BotR.
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]
References
Raffestin, S., Dupuy, B., Marvaud, J. and Popoff, M. (2004). BotR/A and TetR are alternative RNA polymerase sigma factors controlling the expression of the neurotoxin and associated protein genes in Clostridium botulinum type A and Clostridium tetani. Molecular Microbiology, 55(1), pp.235-249.