Difference between revisions of "Part:BBa K3142012"
 
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<partinfo>BBa_K3142012 short</partinfo>
 
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Low glucose environments cause the cAMP-CRP complex to bind target DNA and regulate hundreds of gene targets. [1] There are 3 classes of CRP promoters, categorized by the number and placement of CRP binding sites within each. Class I promoters have one CRP binding site at various distances upstream of the -35 box which interact with the α-C Terminal Domain (α-CTD) of RNA Polymerase (RNAP) . [16]Class II promoters have CRP encompassing the -35 box, and interact with α-CTD and α-NTD of RNAP for promoter recruiting .[ 17] This site partially occludes the σ70 subunit of RNAP, and is likely why multiple interactions with cAMP-CRP are made. Class III promoters contain a mix of multiple Class I and II CRP sites. [16]
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PT-αcrp is a Glucose starved promoter，which is response to glucose starvation.
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Three versions of each promoter were generated: (1) “full length” promoters (crp) starting ~ 500 bp upstream of the open reading frame and ending at the transcription start site (2) minimal “truncated” promoters (T-crp) containing ~ 50 bp upstream of the transcription start site with care taken to include the full CRP binding site, and (3) enhanced truncated promoters (T-αcrp) which include a modified enhancer element from the CC(-41.5)α(-63) synthetic elements upstream of the T-crp promoters. 20
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It can be used to Initiate gene expressing in response to glucose starvation.
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'''what it is'''：Glucose starved promoter
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'''what it does'''：Response to glucose starvation
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'''how to use it in their projects'''：Initiating the autolysis gene expressing lactic acid bacteria in response to glucose starvation
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==SZPT-China==
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===Engineering Success Made by SZU-China 2021===
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To ensure that the transgenic bacteria are not harmful to the environment, we constructed the autolytic enzyme gene acmA of the lactic acid bacteria downstream of the glucose starvation promoter. The results of the construction of the recombinant plasmid pMG36e-pα-crp-acmA named pMG36e-P-a are shown in Fig. 18. The results of LAB MG1363 and LAB MG1363 with pMG36e-p - a after induction of recombinant bacteria by different glucose concentrations are shown in Fig. 18. The results showed that when the concentration of glucose in the environment was less than 0.05%, the growth of recombinant bacteria was significantly inhibited.  
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[[file:p-a.png|center|500px]]
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SZU-China team has added quantitative experimental characterization data to an existing Part from the Registry of Standard Biological Parts and documented the experimental characterization on the Part's Main Page on the Registry.
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This year, We identified a glucose-sensitive promoter pt-αcrp(BBa_K3142012) that can express subsequent genes at low glucose levels to prevent bacteria from escaping to the outside environment. We added the mazF gene, which encodes an endoribonuclease that will be expressed to suicidal effect when glucose levels are low. We verified our system by inducing expression with different glucose concentrations and observing the survival rate of engineered bacteria and supplemented experimental data for the promoter.
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[[File:T--SZU-China--BBa K3838654-GLUK.png|400px|thumb|center|Fig1.Survival curves at different glucose concentrations]]
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As shown in figure 1, compared with the control group, the growth of the bacteria transformed with the plasmids with the glucose-sensitive suicide switch was basically significantly inhibited, and the lower the glucose concentration in the environment, the more significant the inhibition effect was, and the lower the survival rate was reflected in the experiment. The contrast of the line chart shows the difference between the experimental group and the control group. Based on this, we believe that the glucose sensitive kill switch can work normally.
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===Reference===
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*William Henry Bothfeld, Grace Kapov, and Keith Tyo. A glucose-sensing toggle switch for autonomous, high productivity genetic control. ACS Synth. 2017, 6(7):1296-1304.

Latest revision as of 15:04, 18 October 2021


Glucose starvation promoter（PT-αcrp）

PT-αcrp is a Glucose starved promoter，which is response to glucose starvation. It can be used to Initiate gene expressing in response to glucose starvation.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 26
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 26
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 26
    Illegal BamHI site found at 1
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 26
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 26
  • 1000
    COMPATIBLE WITH RFC[1000]


Engineering Success Made by SZU-China 2021

SZU-China team has added quantitative experimental characterization data to an existing Part from the Registry of Standard Biological Parts and documented the experimental characterization on the Part's Main Page on the Registry. This year, We identified a glucose-sensitive promoter pt-αcrp(BBa_K3142012) that can express subsequent genes at low glucose levels to prevent bacteria from escaping to the outside environment. We added the mazF gene, which encodes an endoribonuclease that will be expressed to suicidal effect when glucose levels are low. We verified our system by inducing expression with different glucose concentrations and observing the survival rate of engineered bacteria and supplemented experimental data for the promoter.

Fig1.Survival curves at different glucose concentrations

As shown in figure 1, compared with the control group, the growth of the bacteria transformed with the plasmids with the glucose-sensitive suicide switch was basically significantly inhibited, and the lower the glucose concentration in the environment, the more significant the inhibition effect was, and the lower the survival rate was reflected in the experiment. The contrast of the line chart shows the difference between the experimental group and the control group. Based on this, we believe that the glucose sensitive kill switch can work normally.

Reference

  • William Henry Bothfeld, Grace Kapov, and Keith Tyo. A glucose-sensing toggle switch for autonomous, high productivity genetic control. ACS Synth. 2017, 6(7):1296-1304.