Difference between revisions of "Part:BBa K1153000"

Revision as of 23:48, 16 October 2016

NorV promoter. NOx detector.

Our Biobrick has been designed to enable the detection of NO using the norV gene promoter, cloned from E.coli K12 TOP10 cells.

In the presence of NOx it will promote the downstream genes in the plasmid, producing a protein in response to the environmental stimulus.

Contribution Group: ETH Zurich 2016 Author: Asli Azizoglu Summary: We cloned and characterised the norV promoter, and sent it to the registry as a biobrick.

Promoter norV (PnorV) is the native promoter controlling the nitric oxide reduction operon (norRVW) in E. Coli. [1] It's transcriptional regulator, NorR, can sense nitric oxide and activate gene expression.

We cloned PnorV upstream of super folder GFP for characterisation. This construct was expressed on a medium copy plasmid (p15A). Since E.coli natively produce NorR, we relied on this to activate the promoter. Below is the dose response curve we obtained.

NorV promoter activated by nitric oxide, released from DETA/NO. Above 15uM of DETA/NO effects cell growth and is not included in the dose response.

References: [1] Gardner, A. M. "Regulation Of The Nitric Oxide Reduction Operon (Norrvw) In Escherichia Coli. ROLE OF Norr AND Sigma 54 IN THE NITRIC OXIDE STRESS RESPONSE". Journal of Biological Chemistry 278.12 (2003): 10081-10086.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

@@ Line 9: / Line 9: @@
 '''Group:''' ETH Zurich 2016
 '''Author:''' Asli Azizoglu
-'''Summary:''' We produced and characterised the norV promoter, and sent it to the registry as a biobrick.
+'''Summary:''' We cloned and characterised the norV promoter, and sent it to the registry as a biobrick.
 Promoter norV (PnorV) is the native promoter controlling the nitric oxide reduction operon (norRVW) in E. Coli. [1] It's transcriptional regulator, NorR, can sense nitric oxide and activate gene expression.
+We cloned  PnorV upstream of super folder GFP for characterisation. This construct was expressed on a medium copy plasmid (p15A). Since ''E.coli'' natively produce NorR, we relied on this to activate the promoter. Below is the dose response curve we obtained.
+[[File:PnorVdoseresponse.png|200px|thumb|center|NorV promoter activated by nitric oxide, released from DETA/NO. Above 15uM of DETA/NO effects cell growth and is not included in the dose response.]]
 References: [1] Gardner, A. M. "Regulation Of The Nitric Oxide Reduction Operon (Norrvw) In Escherichia Coli. ROLE OF Norr AND Sigma 54 IN THE NITRIC OXIDE STRESS RESPONSE". Journal of Biological Chemistry 278.12 (2003): 10081-10086.