Difference between revisions of "Part:BBa K4239006"

 
Line 3: Line 3:
 
<partinfo>BBa_K4239006 short</partinfo>
 
<partinfo>BBa_K4239006 short</partinfo>
  
−
aze
+
<h2>Sequence and Features</h2>
 +
<partinfo>BBa_K4239006 SequenceAndFeatures</partinfo>
 +
 
 +
<html>
 +
 
 +
<h2>Description</h2>
 +
 
 +
<p><i>fiatluxC</i> and <i>fiatluxD</i> are made to be used with <i>fiatluxE</i>. They code for a fatty acid reductase. With the subparts coding from <i>fiatluxE</i>, they form a complex that recycles fatty acids to fatty aldehydes. Fatty aldehydes will be used as a substrat for the luciferase protein.</p>
 +
 
 +
<p>The systeme <i>fiatluxC/fiatluxD/fiatluxE</i> is made to be used with <i>fiatluxA</i> and <i>fiatluxB</i>, gathered in the <i>fiatluxCDABE</i> operon. <i>fiatluxA</i> and <i>fiatluxB</i> code for the luciferase protein</p>
 +
 
 +
<p><i>Fiatlux</i> genes come from <i>ilux</i> genes (C, D, A, B, E). They were modified to remove every Igem restriction site (EcoR1, Xba1, Spe1 and Pst1) included in genes. They were also adapted to include the biobrick format.</p>
 +
 
 +
<p>The <i>ilux</i> operon was born from a mutated natural luminescence operon present in the bacteria P.luminescens: the <i>lux</i> operon. These mutations were error-prone PCR induced according to Gregor et al.’s study in 2018 (Gregor et al. 2018). The aim was to create a system of genes that produced more light than the <i>lux</i> system.</p>
 +
 
 +
 
 +
 
 +
<h2>Construction</h2>
 +
 
 +
<p><i>FiatluxCD</i> was directly synthesized with the promoter J23117 <a href="https://parts.igem.org/Part:BBa_J23117" class="pr-0" target="_blank">(BBa_J23117)</a>. It is a constitutive and weak promoter, so no inducer needs to be added. <i>FiatluxCD</i> and their promoter were ordered in the plasmid pACYDuet-1, a p15A low-copy plasmid, and transformed it into <i>E.coli</i> DH5α.</p>
 +
 
 +
<h2>References</h2>
 +
 
 +
<p><i> Gregor C, Gwosch KC, Sahl SJ, Hell SW. Strongly enhanced bacterial bioluminescence with the ilux operon for single-cell imaging. Proc Natl Acad Sci U S A. 2018 Jan 30;115(5):962-967. doi: 10.1073/pnas.1715946115. Epub 2018 Jan 16. PMID: 29339494; PMCID: PMC5798359.</i> </p>
 +
 
  
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here
 
===Usage and Biology===
 
===Usage and Biology===
−
 
−
<!-- -->
 
−
<span class='h3bb'>Sequence and Features</span>
 
−
<partinfo>BBa_K4239006 SequenceAndFeatures</partinfo>
 
  
  

Revision as of 16:20, 8 October 2022


Enhanced luciferase substrate forming subunits fiatluxCD and the promoter J23117

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
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 50
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 1366

Description

fiatluxC and fiatluxD are made to be used with fiatluxE. They code for a fatty acid reductase. With the subparts coding from fiatluxE, they form a complex that recycles fatty acids to fatty aldehydes. Fatty aldehydes will be used as a substrat for the luciferase protein.

The systeme fiatluxC/fiatluxD/fiatluxE is made to be used with fiatluxA and fiatluxB, gathered in the fiatluxCDABE operon. fiatluxA and fiatluxB code for the luciferase protein

Fiatlux genes come from ilux genes (C, D, A, B, E). They were modified to remove every Igem restriction site (EcoR1, Xba1, Spe1 and Pst1) included in genes. They were also adapted to include the biobrick format.

The ilux operon was born from a mutated natural luminescence operon present in the bacteria P.luminescens: the lux operon. These mutations were error-prone PCR induced according to Gregor et al.’s study in 2018 (Gregor et al. 2018). The aim was to create a system of genes that produced more light than the lux system.

Construction

FiatluxCD was directly synthesized with the promoter J23117 (BBa_J23117). It is a constitutive and weak promoter, so no inducer needs to be added. FiatluxCD and their promoter were ordered in the plasmid pACYDuet-1, a p15A low-copy plasmid, and transformed it into E.coli DH5α.

References

Gregor C, Gwosch KC, Sahl SJ, Hell SW. Strongly enhanced bacterial bioluminescence with the ilux operon for single-cell imaging. Proc Natl Acad Sci U S A. 2018 Jan 30;115(5):962-967. doi: 10.1073/pnas.1715946115. Epub 2018 Jan 16. PMID: 29339494; PMCID: PMC5798359.