Difference between revisions of "Part:BBa K2442202"

Revision as of 22:32, 1 November 2017

Ptet + RBS + MtlR

This part contains the mtlR coding sequence along with the ptet promoter. mtlR coding sequence is derived from the mannitol operon from Pseudomonas Fluorescens. The ptet promoter is the well characterised part R0040. This Biobrick was used in conjunction with BBa_K2442203. By splitting the regulatory protein (mtlR) from the promoter it regulates (pmtle) allowed for greater control over the expression of GFP.

Usage and Biology

The mtlR regulatory protein is found naturally in Pseudomonas fluorescens. The mtlR protein has been described as a multi-sugar transcriptional regulator. There is some evidence to suggest that the protein functions as a dimer. Naturally this multi-sugar regulator functions to switch on sugar metabolism genes in P.Flourescens in the presence of organic sugars.

The aim of our project was to harness the natural regulatory abilities of the mtlR protein and use this alongside the promoter that it regulates (pmtle). We also aimed to combine this regulatory system with a reporter construct BBa_K2442203 to evoke GFP response. We used the sequence of the P.Flourescens mtlR gene to construct a coding sequence that was regulated upstream by the ptet promoter. By cotrasnforming E.coli cells with BBa_K2442202 and BBa_K2442203 we were able to create a sugar responsive assay that produced GFP via induction by sugars.

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 3: / Line 3: @@
 <partinfo>BBa_K2442202 short</partinfo>
-This part contains the
+This part contains the mtlR coding sequence along with the ptet promoter. mtlR coding sequence is derived from the mannitol operon from Pseudomonas Fluorescens. The ptet promoter is the well characterised part [https://parts.igem.org/Part:BBa_R0040 R0040].
+This Biobrick was used in conjunction with BBa_K2442203. By splitting the regulatory protein (mtlR) from the promoter it regulates (pmtle) allowed for greater control over the expression of GFP.
-This part contains the ptet promoter (R0040) with a strong ribosome binding site (B0032).
-These components lie upstream of the protein coding sequence for the mtlR protein native to <i>Pseudomonas fluorescens</i>.
+===Usage and Biology===
-This part was intended to be used as regulator construct in our system. The mtlR protein is documented to regulate the activity of the native promoter to Pseudomonas , mtle. Currently it is thought that sugars can induce the dimerisation of the mtlR protein and this complex can in turn activate pmtle. This system can be coupled to GFP to create a reporter construct.
+The mtlR regulatory protein is found naturally in Pseudomonas fluorescens. The mtlR protein has been described as a multi-sugar transcriptional regulator. There is some evidence to suggest that the protein functions as a dimer. Naturally this multi-sugar regulator functions to switch on sugar metabolism genes in P.Flourescens in the presence of organic sugars.
-We tested the regulatory properties of this part by transforming <i>E. coli</i> with the reporter plasmid K2442206. <i>Figure 1</i> illustrates the result of this and indicates the regulatory property of this part works.
-https://static.igem.org/mediawiki/parts/a/a5/T-Glasgow-finalmtlrdiagram.png
+The aim of our project was to harness the natural regulatory abilities of the mtlR protein and use this alongside the promoter that it regulates (pmtle). We also aimed to combine this regulatory system with a reporter construct BBa_K2442203 to evoke GFP response.
+We used the sequence of the P.Flourescens mtlR gene to construct a coding sequence that was regulated upstream by the ptet promoter. By cotrasnforming E.coli cells with BBa_K2442202 and BBa_K2442203 we were able to create a sugar responsive assay that produced GFP via induction by sugars.
-<i>Figure 1</i> The Regulator (K2442202) + WT reporter construct evoked GFP production with induction achieved by various sugars. In particular when ribose and sorbitol were used as inducers.
-===Usage and Biology===