Difference between revisions of "Part:BBa K3411020"

Revision as of 23:37, 27 October 2020

NarP Nitrate/Nitrite Sensor

Lambert_GA iGEM 2020's novel NarP system, BBa_K3411020, is also modeled after E.coli’s natural nitrite sensor in the NAR Operon. Because the NarL system (BBa_K1682018) detects nitrate, Lambert iGEM needed a method to detect levels of nitrite: NarP, which detects both nitrate and nitrite. By calculating the corrected difference, the system would have concentrations for both nutrients. Lambert iGEM’s novel NarP composite part BBa_K3411020 and the NarP gene BBa_K3411010 are also modeled after E.coli’s natural nitrite sensor. Similarly to NarL, the promoter BBa_J23106, a constitutive Anderson Promoter, produces TetR, which represses the PtetO promoter, producing NarP. By regulating the amount of aTc, the molecule that inhibits TetR, the lab can control NarP’s expression, the product of the PtetO promoter. NarQ, the native membrane-bound protein, senses the amount of nitrate and/or nitrite in the cell: in the presence of nitrite and/or nitrate, NarQ will phosphorylate NarP, activating it, but if there is no nitrite and/or nitrate present, NarQ will not phosphorylate NarP [1]. If NarP is phosphorylated, it will activate the nirB promoter, producing GFP [2]. Lambert iGEM plans to correlate GFP levels to nitrite through mathematical modeling.

Lambert_GA iGEM 2020's plans on cloning and characterizing our NarP composite part in the coming year as a part of our two-year project, AgroSense.

References

[1] Darwin A.J., & Stewart V. (1996) The NAR Modulon Systems: Nitrate and Nitrite Regulation of Anaerobic Gene Expression. In: Regulation of Gene Expression in Escherichia coli. Springer, Boston, MA. https://doi.org/10.1007/978-1-4684-8601-8_17 [2] Wang H., & Gunsalus R. (2000). The nrfA and nirB Nitrite Reductase Operons in Escherichia coli Are Expressed Differently in Response to Nitrate than to Nitrite. Journal of Bacteriology, 182(20): 5813–5822. doi: 10.1128/jb.182.20.5813-5822.2000

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 2000
Illegal NheI site found at 2023
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal NgoMIV site found at 2727
Illegal AgeI site found at 1371
1000
COMPATIBLE WITH RFC[1000]

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 <partinfo>BBa_K3411020 short</partinfo>
-Lambert_GA iGEM 2020's novel NarP system, [https://parts.igem.org/Part:BBa_K3411020 BBa_K3411020], is also modeled after <i>E.coli</i>’s natural nitrite sensor in the NAR Operon. Similar to NarL, the promoter [https://parts.igem.org/Part:BBa_J23106 BBa_J23106], a constitutive Anderson Promoter, produces TetR, which represses the PtetO promoter, producing NarP. By regulating the amount of aTc, the molecule that inhibits TetR, the lab can control NarP’s expression, the product of the PtetO promoter. NarQ, the native membrane-bound protein, senses the amount of nitrate and/or nitrite in the cell: In the presence of nitrite and/or nitrate, NarQ will phosphorylate NarP, activating it, but if there is no nitrite and/or nitrate present, NarQ will not phosphorylate NarP. If NarP is phosphorylated, it will activate the nirB promoter, producing GFP.
+Lambert_GA iGEM 2020's novel NarP system, [https://parts.igem.org/Part:BBa_K3411020 BBa_K3411020], is also modeled after <i>E.coli</i>’s natural nitrite sensor in the NAR Operon. Because the NarL system ([https://parts.igem.org/Part:BBa_K1682018 BBa_K1682018]) detects nitrate, Lambert iGEM needed a method to detect levels of nitrite: NarP, which detects both nitrate and nitrite. By calculating the corrected difference, the system would have concentrations for both nutrients. Lambert iGEM’s novel NarP composite part BBa_K3411020 and the NarP gene BBa_K3411010 are also modeled after E.coli’s natural nitrite sensor. Similarly to NarL, the promoter BBa_J23106, a constitutive Anderson Promoter, produces TetR, which represses the PtetO promoter, producing NarP. By regulating the amount of aTc, the molecule that inhibits TetR, the lab can control NarP’s expression, the product of the PtetO promoter. NarQ, the native membrane-bound protein, senses the amount of nitrate and/or nitrite in the cell: in the presence of nitrite and/or nitrate, NarQ will phosphorylate NarP, activating it, but if there is no nitrite and/or nitrate present, NarQ will not phosphorylate NarP [1]. If NarP is phosphorylated, it will activate the nirB promoter, producing GFP [2]. Lambert iGEM plans to correlate GFP levels to nitrite through mathematical modeling.
 Lambert_GA iGEM 2020's plans on cloning and characterizing our NarP composite part in the coming year as a part of our two-year project, AgroSense.
+===References===
+[1] Darwin A.J., & Stewart V. (1996) The NAR Modulon Systems: Nitrate and Nitrite Regulation of Anaerobic Gene Expression. In: Regulation of Gene Expression in Escherichia coli. Springer, Boston, MA. https://doi.org/10.1007/978-1-4684-8601-8_17
+[2] Wang H., & Gunsalus R. (2000). The nrfA and nirB Nitrite Reductase Operons in Escherichia coli Are Expressed Differently in Response to Nitrate than to Nitrite. Journal of Bacteriology, 182(20): 5813–5822. doi: 10.1128/jb.182.20.5813-5822.2000
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