Difference between revisions of "Part:BBa K1067006"
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===Characterization=== | ===Characterization=== | ||
| − | To characterize this biobrick, we ran an anaerobic experiment in which we added a known amount of nitrite to an anaerobic flask, and measured the nitrous oxide produced using a microsensor. We had a lot of trouble with the microsensor and were not able to conclusively prove that we were producing nitrous oxide. | + | To characterize this biobrick, we ran an anaerobic experiment in which we added a known amount of nitrite to an anaerobic flask, and measured the nitrous oxide produced using a microsensor. We had a lot of trouble with the microsensor (see especially hour 4 on the below graph) and were not able to conclusively prove that we were producing nitrous oxide. We would recommend rerunning the experiment with a more reliable sensor. |
| − | [[File:dtu-nir-char.png]] | + | [[File:dtu-nir-char.png|300px|Nitrous oxide produced over time, measured with a microsensor in an anaerobic environment. The times that nitrite was added is indicated by the vertical lines. The data does not indicate that cells transformed with this biobrick (BBa_K1067006) show a strong response to nitrite.]] |
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Revision as of 13:56, 12 October 2013
Denitrification operon from Pseudomonas aeruginosa
This biobrick is the whole denitrification region from Pseudomonas aeruginosa PAO1 excluding only the enzyme in last step; Nitrous-oxide reductase ([http://ecocyc.org/PAER941193/NEW-IMAGE?type=LOCUS-POSITION&object=GLIQ-518&chromosome=CHROMOSOME-1 link to Biocyc]).
Usage and Biology
This brick has been shown hard to work with because it is a large region, all under one promoter. We purpose that splitting it into the 3 operons; like the conformation natively in P. aeruginosa and adding a separate promoter for each would make it easier to work with. If trying to to control the whole region under the same promoter one might experience premature mRNA transcripts because RNAP will fall of before the end of the region.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 3445
Illegal PstI site found at 6521
Illegal PstI site found at 7162 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 3445
Illegal PstI site found at 6521
Illegal PstI site found at 7162
Illegal NotI site found at 4536
Illegal NotI site found at 6121 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 3445
Illegal BglII site found at 197
Illegal BglII site found at 2942
Illegal BglII site found at 3335
Illegal XhoI site found at 6639 - 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 3445
Illegal PstI site found at 6521
Illegal PstI site found at 7162 - 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 3445
Illegal PstI site found at 6521
Illegal PstI site found at 7162
Illegal NgoMIV site found at 799
Illegal NgoMIV site found at 1926
Illegal NgoMIV site found at 1935
Illegal NgoMIV site found at 2357
Illegal NgoMIV site found at 2381
Illegal NgoMIV site found at 2467
Illegal NgoMIV site found at 3292
Illegal AgeI site found at 1663
Illegal AgeI site found at 6506 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 841
Illegal BsaI.rc site found at 973
Illegal BsaI.rc site found at 1706
Illegal BsaI.rc site found at 3391
Illegal BsaI.rc site found at 3693
Illegal BsaI.rc site found at 5125
Illegal BsaI.rc site found at 5676
Illegal SapI site found at 6182
Illegal SapI site found at 7434
Characterization
To characterize this biobrick, we ran an anaerobic experiment in which we added a known amount of nitrite to an anaerobic flask, and measured the nitrous oxide produced using a microsensor. We had a lot of trouble with the microsensor (see especially hour 4 on the below graph) and were not able to conclusively prove that we were producing nitrous oxide. We would recommend rerunning the experiment with a more reliable sensor.
