Difference between revisions of "Part:BBa K2556333"
 
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Under dark conditions, the phosphate group is transferred from Yf1 protein to FixJ protein, the phosphorylated FixJ protein activates the PfixK2 promoter and then activates the downstream gene expression. When induced by light, the phosphorylation of Fixj protein will be blocked, and the expression of genes regulated by PfixK2 promoters will be inhibited.
 
Under dark conditions, the phosphate group is transferred from Yf1 protein to FixJ protein, the phosphorylated FixJ protein activates the PfixK2 promoter and then activates the downstream gene expression. When induced by light, the phosphorylation of Fixj protein will be blocked, and the expression of genes regulated by PfixK2 promoters will be inhibited.
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https://static.igem.org/mediawiki/2018/4/43/T--ZJUT-China--part101.png
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https://static.igem.org/mediawiki/2018/9/99/T--ZJUT-China--101702.png
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<h1>Characterize</h1>
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===<h1>Characterize</h1>===
 
<p>To further verify the effectiveness of the system, the plasmids with the system were transferred into different hosts and cultured for more than 24 hours under blue light and dark light respectively. The fluorescence value of eGFP in the culture medium was measured.
 
<p>To further verify the effectiveness of the system, the plasmids with the system were transferred into different hosts and cultured for more than 24 hours under blue light and dark light respectively. The fluorescence value of eGFP in the culture medium was measured.
 
</p>
 
</p>
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<h1>Experimental Results</h1>
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===<h1>Experimental Results</h1>===
 
<p>Detection of expression efficiency of primary light control system in different hosts</p>
 
<p>Detection of expression efficiency of primary light control system in different hosts</p>
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<html>
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<img src="https://static.igem.org/mediawiki/2018/2/29/T--ZJUT-China--designyu7.png" alt="" width="600px">
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<div class="note">Figure 1. expression efficiency in DH5α</div>
 
<div class="note">Figure 1. expression efficiency in DH5α</div>
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https://static.igem.org/mediawiki/2018/f/f2/T--ZJUT-China--designyu8.png
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<img src="https://static.igem.org/mediawiki/2018/f/f2/T--ZJUT-China--designyu8.png" alt="" width="600px">
 
<p>Figure 2. expression efficiency in BL21</p>
 
<p>Figure 2. expression efficiency in BL21</p>
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https://static.igem.org/mediawiki/2018/d/de/T--ZJUT-China--designyu9.png
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<img src="https://static.igem.org/mediawiki/2018/9/9d/T--ZJUT-China--part3.png" alt="" width="600px">
 
<p>Figure 3. expression efficiency in 4MG1655</p>
 
<p>Figure 3. expression efficiency in 4MG1655</p>
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http://2018.igem.org/File:T--ZJUT-China--designyu10.png
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<img src="https://static.igem.org/mediawiki/2018/9/9e/T--ZJUT-China--designyu10.png" alt="" width="600px">
 
<p>Figure 4. expression efficiency in BW25113</p>
 
<p>Figure 4. expression efficiency in BW25113</p>
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<p>The dusk-eGFP-pUC57 plasmid was transferred into the host of DH5α,BL21,4MG1655 and BW25113, cultured under dark or blue light for a period of time and the fluorescence value of eGFP was detected. After 24 hours, the fluorescence value of the experimental group in the dark culture showed an obvious increasing trend. There was no significant increase in fluorescence value in both the experimental group under blue light and the negative control group.  
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</html>
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<p>The dusk-eGFP-pUC57 plasmid was transferred into the host of DH5α,BL21,4MG1655 and BW25113, cultured under dark or blue light for a period of time and the fluorescence value of eGFP was detected. After 24 hours, the fluorescence value of the experimental group in the dark culture showed an obvious increasing trend. There was no significant increase in fluorescence value in both the experimental group under blue light and the negative control group.
 
</p>
 
</p>
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<br>  <br>
 
<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
 
<partinfo>BBa_K2556333 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K2556333 SequenceAndFeatures</partinfo>
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===<h1>References</h1>===
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<p>
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[1]Wang G, Lu X, Zhu Y, et al. A light-controlled cell lysis system in bacteria.[J]. Journal of Industrial Microbiology & Biotechnology, 2018:1-4.
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<br>[2]Wu H, Wang Y, Wang Y, et al. Quantitatively relating gene expression to light intensity via the serial connection of blue light sensor and CRISPRi[J]. Acs Synthetic Biology, 2014, 3(12):979.
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<br>[3]Gardner L, Deiters A. Light-Controlled Synthetic Gene Circuits[J]. Current Opinion in Chemical Biology, 2012, 16(3-4):292-299.
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</p>
  
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<!-- Uncomment this to enable Functional Parameter display
 
 
===Functional Parameters===
 
===Functional Parameters===
 
<partinfo>BBa_K2556333 parameters</partinfo>
 
<partinfo>BBa_K2556333 parameters</partinfo>
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Latest revision as of 19:31, 17 October 2018


Light control system with eGFP


Under dark conditions, the phosphate group is transferred from Yf1 protein to FixJ protein, the phosphorylated FixJ protein activates the PfixK2 promoter and then activates the downstream gene expression. When induced by light, the phosphorylation of Fixj protein will be blocked, and the expression of genes regulated by PfixK2 promoters will be inhibited. T--ZJUT-China--101702.png

Characterize

To further verify the effectiveness of the system, the plasmids with the system were transferred into different hosts and cultured for more than 24 hours under blue light and dark light respectively. The fluorescence value of eGFP in the culture medium was measured.

Experimental Results

Detection of expression efficiency of primary light control system in different hosts

Figure 1. expression efficiency in DH5α

Figure 2. expression efficiency in BL21

Figure 3. expression efficiency in 4MG1655

Figure 4. expression efficiency in BW25113

The dusk-eGFP-pUC57 plasmid was transferred into the host of DH5α,BL21,4MG1655 and BW25113, cultured under dark or blue light for a period of time and the fluorescence value of eGFP was detected. After 24 hours, the fluorescence value of the experimental group in the dark culture showed an obvious increasing trend. There was no significant increase in fluorescence value in both the experimental group under blue light and the negative control group.



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 NgoMIV site found at 605
    Illegal NgoMIV site found at 677
    Illegal NgoMIV site found at 767
    Illegal NgoMIV site found at 785
    Illegal NgoMIV site found at 1297
    Illegal NgoMIV site found at 1590
    Illegal NgoMIV site found at 1684
    Illegal AgeI site found at 319
    Illegal AgeI site found at 1465
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1354
    Illegal BsaI.rc site found at 218

References

[1]Wang G, Lu X, Zhu Y, et al. A light-controlled cell lysis system in bacteria.[J]. Journal of Industrial Microbiology & Biotechnology, 2018:1-4.
[2]Wu H, Wang Y, Wang Y, et al. Quantitatively relating gene expression to light intensity via the serial connection of blue light sensor and CRISPRi[J]. Acs Synthetic Biology, 2014, 3(12):979.
[3]Gardner L, Deiters A. Light-Controlled Synthetic Gene Circuits[J]. Current Opinion in Chemical Biology, 2012, 16(3-4):292-299.

Functional Parameters