Difference between revisions of "Part:BBa K4645019"

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===Usage and Biology===
 
===Usage and Biology===
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<p>J23100-B0030-luxR-B0017-Plux-tetR-T7 terminator-Ptet-amcyan- T7 terminator
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<p>
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We transformed this biobrick into E. coli to regulate expression of genes. With the growing of bacteria, AHL accumulates. While the concentration of AHL reached the threshold value, the expression of NeuA, NeuB and TetR would be activated. Then TetR shut down the expression of amcyan in downstream.
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We transformed this biobrick into <i>E. coli</i> BL21(DE3) to regulate the expression of downstream genes. With the growing of bacteria, AHL accumulates. While the concentration of AHL reached the threshold value, the expression of NeuA, NeuB and TetR would be activated. Then TetR shut down the expression of AmCyan in downstream.
 
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<title>What？</title>
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<center><img src="https://static.igem.wiki/teams/4645/wiki/quorum-sensing/luxr-tetr-us.jpg" style="width:65%; "></center>
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<center><b>Figure 1. The composite part we assembled. </b></center>
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===Characterization===
 
===Characterization===
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<p>Blank BL21, BL21 transformed with this biobrick, were cultured in microplate reader 37°C, 220 rpm for 10 hours and detected OD600, fluorescence intensity (458 nm excitation light, 489 nm emission light) every 10 minutes.</p>
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<p>Blank <i>E.coli</i> BL21(DE3), engineered <i>E.coli</i> BL21(DE3) with this biobrick were cultured in microplate reader 37°C, 220 rpm for 10 hours and detected OD<sub>600</sub>, fluorescence intensity (458 nm excitation light, 489 nm emission light) every 10 minutes.</p>
 
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<center><img src=" https://static.igem.wiki/teams/4645/wiki/quorum-sensing/luxr-tetr-chara.jpg" style="width:50%; "></center>
 
<center><img src=" https://static.igem.wiki/teams/4645/wiki/quorum-sensing/luxr-tetr-chara.jpg" style="width:50%; "></center>
 
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<center><b>Figure 2. Changes of fluorescence intensity / OD600 over time.</b></center>
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<center><b>Figure 2. Changes of fluorescence intensity / OD<sub>600</sub> in blank <i>E.coli</i> BL21(DE3), engineered <i>E.coli</i> BL21(DE3): <i>luxR-tetR</i> over time.</b></center>
 
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Latest revision as of 14:59, 12 October 2023


J23100-B0030-luxR-B0017-lux pR-tetR-B0015-PTet-amcyan-B0017

Verification of Not-Gate.


Usage and Biology

We transformed this biobrick into E. coli BL21(DE3) to regulate the expression of downstream genes. With the growing of bacteria, AHL accumulates. While the concentration of AHL reached the threshold value, the expression of NeuA, NeuB and TetR would be activated. Then TetR shut down the expression of AmCyan in downstream.

What？


Figure 1. The composite part we assembled.

Characterization

Blank E.coli BL21(DE3), engineered E.coli BL21(DE3) with this biobrick were cultured in microplate reader 37°C, 220 rpm for 10 hours and detected OD600, fluorescence intensity (458 nm excitation light, 489 nm emission light) every 10 minutes.

What？


Figure 2. Changes of fluorescence intensity / OD600 in blank E.coli BL21(DE3), engineered E.coli BL21(DE3): luxR-tetR over time.


The result shows that in the early growth stage, transformed bacteria expressed amcyan fluorescent protein continually. When the group density reached the threshold value, the expression of amcyan fluorescent protein began to be blocked up.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 2541
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 1027