Difference between revisions of "Part:BBa K3725120"

Latest revision as of 20:34, 17 December 2021

PhoB Mutated Consensus Promoter with GFP Reporter

DESIGN

The promoter BBa_K3725130 was designed by replacing parts of BBa_K116401 with the PhoB consensus sequence. A consensus sequence is a calculated order of the most repeated nucleotides found at each position in an alignment of multiple other sequences that have a similar function. By replacing the downstream Pho Regulon with GFP, the part was expected to express fluorescence in the presence of low levels of extracellular phosphate.

CHARACTERIZATION

Utilizing the phosphate range of 0-150μM and characterization protocol (See: Modified Protocol), we characterized the BBa_K3725120 phosphate sensor (Figure 1). However, after overnight incubation, we noticed that the cells produced no fluorescence. Despite resuspending the cells in MOPS media, which contains relatively less phosphate than LB, the part did not exhibit greater fluorescence. Since we confirmed the cloning of the part with successful sequencing results, we concluded that the part was not efficient as a phosphate sensor and discontinued characterization.

Figure 1. Characterization curve for BBa_K3725120 for phosphate concentrations between 0μM and 150μM.

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
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 1148

@@ Line 4: / Line 4: @@
 ====DESIGN====
-The promoter BBa_K3725130 was designed by replacing parts of BBa_K116401 with the PhoB consensus sequence. A consensus sequence is a calculated order of the most repeated nucleotides found at each position in an alignment of multiple other sequences that have a similar function. By replacing the downstream Pho Regulon with GFP, the part was expected to express fluorescence in the presence of low levels of extracellular phosphate.
+The promoter [https://parts.igem.org/Part:BBa_K3725130 BBa_K3725130] was designed by replacing parts of [https://parts.igem.org/Part:BBa_K116401 BBa_K116401] with the PhoB consensus sequence. A consensus sequence is a calculated order of the most repeated nucleotides found at each position in an alignment of multiple other sequences that have a similar function. By replacing the downstream Pho Regulon with GFP, the part was expected to express fluorescence in the presence of low levels of extracellular phosphate.
-====EXPERIENCE====
+====CHARACTERIZATION====
-Utilizing the modified phosphate range and characterization protocol (See: Modified Protocol under Pho), we characterized the BBa_K3725120 phosphate sensor. However, after overnight incubation, we noticed that the cells produced no fluorescence. Despite resuspending the cells in MOPS media, which contains relatively less phosphate than LB, the part did not exhibit greater fluorescence. Since we confirmed the cloning of the part with successful sequencing results, we concluded that the part was not efficient as a phosphate sensor and discontinued characterization.
+Utilizing the phosphate range of 0-150μM and characterization protocol (See: [https://2021.igem.org/Team:Lambert_GA/Pho#:~:text=Engineering%20Success%20page.-,Characterization%20Protocol,-The%20failed%20characterization Modified Protocol]), we characterized the BBa_K3725120 phosphate sensor (Figure 1). However, after overnight incubation, we noticed that the cells produced no fluorescence. Despite resuspending the cells in MOPS media, which contains relatively less phosphate than LB, the part did not exhibit greater fluorescence. Since we confirmed the cloning of the part with successful sequencing results, we concluded that the part was not efficient as a phosphate sensor and discontinued characterization.
+<br>
+[[File:Fjasksdanfnkcnfkncjak.png|thumb|center|800px|Figure 1. Characterization curve for BBa_K3725120 for phosphate concentrations between 0μM and 150μM. ]]
 <!-- Add more about the biology of this part here