Difference between revisions of "Part:BBa K4325020"

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<partinfo>BBa_K4325020 short</partinfo>
 
<partinfo>BBa_K4325020 short</partinfo>
 
===Description===
 
===Description===
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This composite part is a generator consisting of pDawn(cI-LVA)(<partinfo>BBa_K1075044</partinfo>) and X174E(<partinfo>BBa_K1835500</partinfo>).
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This composite part is a generator consisting of pDawn(cI-LVA)(<partinfo>BBa_K1075044</partinfo>), RBS070 (<partinfo>BBa_K4325002</partinfo>), X174E(<partinfo>BBa_K1835500</partinfo>) and T1 terminator(<partinfo>BBa_K3033016</partinfo>).
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[[File:K20 1.png|600px|thumb|center|Figure 1:Genextic circuit of pDawn(cI-LVA)-RBS070-X174E-T1]]
 
===Usage===
 
===Usage===
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<p>The pDawn (cI-LVA)(<partinfo>BBa_K1075044</partinfo>) blue light response system and the lysis gene X174E(<partinfo>BBa_K1835500</partinfo>) were inserted into the pSEVA331 expression vector,which wsa inserted into the <i>E.coli</i> TOP10,screened out bacterial colony that grew in the dark but did not grow under blue light. Finally, the pSEVA331-pDawn-RBS070-X174E-T1 plasmid was selected and inserted into <i> G. hansenii</i> ATCC53582 to verify the responsiveness of pDawn(cI-LVA) to blue light.</p>
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<p>The pDawn (cI-LVA)(<partinfo>BBa_K1075044</partinfo>) blue light responsive system and the lysis gene X174E(<partinfo>BBa_K1835500</partinfo>) were inserted into the pSEVA331 expression vector, which was incorporated into the <i>E.coli</i> TOP10, screened for the bacterial colonies that grew in the dark but did not grow under blue light..</p>
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[[File:K20 1.png|600px|thumb|center|Figure 1: <p></p>Gene circuit of pDawn(cI-LVA)-RBS070-X174E-T1]]
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===Sequence and Features===
 
===Sequence and Features===
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=2022 SZPT-China=
 
=2022 SZPT-China=
 
<h3>Characterization</h3>
 
<h3>Characterization</h3>
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<h4>1.Batch screening of pSEVA331-pDawn(cI-LVA)-RBS070-X174E-T1 in response to blue light lysis in <i>E. coli</i>.</h4>
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<h4>1.Batch screening of pDawn-RBS070-X174E-T1 by evaluating the response to blue light. </h4>
 +
 
 +
<p>As shown in Figure 2, the 11<sup>th </sup> and 15<sup>th </sup> bacterial colonies did not grow under blue light but grow in the dark. In order to further explore the lysis effect of pDawn(cI-LVA)-RBS070-X174E-T1, we tracked the OD<sub>600</sub> value of pDawn(cI-LVA)-RBS070-X174E-T1 and plotted the growth curve diagram. As shown in Figure 3, the lysis effect of pDawn(cI-LVA)-RBS070-X174E-T1 is not well.</p>
 +
[[File:K20 2.png|600px|thumb|center|Figure 2: The growth condition of <i>E. coli</i> TOP10-pSEVA331-pDawn(cI-LVA)-RBS070-X174E-T1 in the dark and under blue light.]]
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[[File:K20 3.png|600px|thumb|center|Figure 3: Growth curve diagram of <i>E. coli</i> TOP10-pSEVA331-pDawn(cI-LVA)-RBS070-X174E-T1.]]
  
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<p>As shown in Figure 2, the 11<sup>th </sup> and 15 <sup>th </sup> bacterial colony did not grow under blue light but  grow in dark. In order to further explore the lysis effect of pSEVA331-pDawn (cI-LVA)-RBS070-X174E-T1-TOP10, we measured the OD<sub>600</sub> value of pSEVA331-pDawn (cI-LVA)-RBS070-X174E-T1-TOP10 periodically andploting the growth curve diagram.As shown in Figure 3, the lysis effect of pSEVA331-pDawn (cI-LVA)-RBS070-X174E-T1-TOP10 is not well.</p>
 
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[[File:K20 2.png|600px|thumb|center|Figure 2: <p></p>The growth condition of pDawn(cI-LVA)-RBS070-X174E-T1-TOP10 in the dark and under the blue light.]]
 
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[[File:K20 3.png|600px|thumb|center|Figure 3: <p></p>Growth curve diagram of pDawn(cI-LVA)-RBS070-X174E-T1-TOP10.]]
 
  
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<h3>2.pSEVA331-pDawn(cI-LVA)-RBS070-X174E-T1 in response to blue photolysis in <i>G. hansenii</i> ATCC53582.</h3>
 
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<p>pDawn(cI-LVA)-RBS070-X174E-T1 was inserted into <i>G. hansenii</i>  ATCC53582 by electroporation and identified. As shown in Figure 4, pDawn(cI-LVA)-RBS070-X174E-T1 was successfully inserted into <i>G. hansenii</i> ATCC53582.</p>
 
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[[File:K26 44.png|600px|thumb|center|Figure 3: <p></p>Growth curve diagram of pDawn(cI-LVA)-RBS070-X174E-T1-TOP10.]]
 
 
<h3>References</h3>
 
<h3>References</h3>
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<p>[1]Robert Ohlendorf, Roee R. Vidavski, Avigdor Eldar et.al.From Dusk till Dawn: One-Plasmid Systems for Light-Regulated Gene Expression.Journal of Molecular Biology,08 Jan 2012, 416(4):534-542.</p>
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<p>[1] Robert Ohlendorf, Roee R. Vidavski, Avigdor Eldar et.al.From Dusk till Dawn: One-Plasmid Systems for Light-Regulated Gene Expression.Journal of Molecular Biology,08 Jan 2012, 416(4):534-542.</p>
 +
<p>[2] Witte, A., Wanner, G., Sulzner, M. & Lubitz, W. Dynamics of PhiX174 protein E-mediated lysis of <i>Escherichia coli.</i> Arch. Microbiol. 157, 381-8 (1992).</p>

Latest revision as of 14:35, 12 October 2022

pDawn-RBS070-X174E-T1

Description

This composite part is a generator consisting of pDawn(cI-LVA)(BBa_K1075044), RBS070 (BBa_K4325002), X174E(BBa_K1835500) and T1 terminator(BBa_K3033016).

Figure 1:Genextic circuit of pDawn(cI-LVA)-RBS070-X174E-T1

Usage

The pDawn (cI-LVA)(BBa_K1075044) blue light responsive system and the lysis gene X174E(BBa_K1835500) were inserted into the pSEVA331 expression vector, which was incorporated into the E.coli TOP10, screened for the bacterial colonies that grew in the dark but did not grow under blue light..


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 2171
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 63
    Illegal NgoMIV site found at 195
    Illegal NgoMIV site found at 289
    Illegal NgoMIV site found at 582
    Illegal NgoMIV site found at 1076
    Illegal NgoMIV site found at 1094
    Illegal NgoMIV site found at 1184
    Illegal AgeI site found at 414
    Illegal AgeI site found at 1542
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1643
    Illegal BsaI.rc site found at 525


2022 SZPT-China

Characterization

1.Batch screening of pDawn-RBS070-X174E-T1 by evaluating the response to blue light.

As shown in Figure 2, the 11th and 15th bacterial colonies did not grow under blue light but grow in the dark. In order to further explore the lysis effect of pDawn(cI-LVA)-RBS070-X174E-T1, we tracked the OD600 value of pDawn(cI-LVA)-RBS070-X174E-T1 and plotted the growth curve diagram. As shown in Figure 3, the lysis effect of pDawn(cI-LVA)-RBS070-X174E-T1 is not well.

Figure 2: The growth condition of E. coli TOP10-pSEVA331-pDawn(cI-LVA)-RBS070-X174E-T1 in the dark and under blue light.
Figure 3: Growth curve diagram of E. coli TOP10-pSEVA331-pDawn(cI-LVA)-RBS070-X174E-T1.


References

[1] Robert Ohlendorf, Roee R. Vidavski, Avigdor Eldar et.al.From Dusk till Dawn: One-Plasmid Systems for Light-Regulated Gene Expression.Journal of Molecular Biology,08 Jan 2012, 416(4):534-542.

[2] Witte, A., Wanner, G., Sulzner, M. & Lubitz, W. Dynamics of PhiX174 protein E-mediated lysis of Escherichia coli. Arch. Microbiol. 157, 381-8 (1992).