Difference between revisions of "Part:BBa K4325024"
 
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<partinfo>BBa_K4325024 short</partinfo>
 
<partinfo>BBa_K4325024 short</partinfo>
 
===Description===
 
===Description===
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This composite part is a generator consisting of pDawn (without LVA tag) (<partinfo>BBa_K4325010 </partinfo>) and X174E (<partinfo>BBa_K1835500 </partinfo>).
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This composite part is a generator consisting of pDawn(cI) (<partinfo>BBa_K4325010 </partinfo>),  X174E (<partinfo>BBa_K1835500 </partinfo>) and T1 terminator (<partinfo>BBa_K3033016 </partinfo>).
 
===Usage===
 
===Usage===
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<p>We inserted the pDawn (without LVA tag) blue light response system (<partinfo>BBa_K4325010 </partinfo>)with the lysis gene X174E (<partinfo>BBa_K1835500 </partinfo>)into the pSEVA331 expression vector, which was inserted into <i> E. coli</i> TOP10. The bacterial colony that grew in the dark but did not grow under blue light were screened to verify the responsiveness of pDawn (without LVA tag) to blue light.
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<p>We inserted the blue light responsive system pDawn(cI) (<partinfo>BBa_K4325010 </partinfo>) and the lysis gene X174E (<partinfo>BBa_K1835500 </partinfo>) into the pSEVA331 expression vector, which was introduced into <i>E. coli</i> TOP10. The bacterial colonies that grew in the dark but did not grow under blue light were screened to verify the responsiveness of pDawn(cI) to blue light.</p>
  
  
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<h3>Characterization</h3>
 
<h3>Characterization</h3>
 
<h4>
 
<h4>
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1.Batch screening of pDawn(without LVA tag)-X174E-T1 in response to blue light lysis in <i> E. coli</i>.</h4>
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1.Batch screening of pDawn(cl)-X174E-T1 by evaluating the response to blue light.</h4>
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[[File:2222222222222222.png|600px|thumb|center|Figure1. (1)pSEVA331-pDawn(cI-LVA)-X174E-LVA-T1; (2)pSEVA331-pDawn(cI)-X174E-T1; (3)pSEVA331-pDawn(cI-LVA)-RBS070-LKD-LVA-T1; (4)pSEVA331-pDawn(cI)-RBS070-LKD-T1 were constructed successfully.]]<P></P> [[File:3333333.png|600px|thumb|center|Figure2. Growth condition of pSEVA331-pDawn(cI)-X174E-T1-TOP10 in dark and under light.]]<p></p> As shown in Figure 1, we successfully constructed pSEVA331-pDawn(cI)-X174E-T1 and transformed it into <i> E. coli</i> TOP10, selection a monoclonal colony and culturing in different growth conditions. As shown in Figure 2, the 9th colony of pSEVA331-pDawn(cI)-X174E-T1-TOP10 grew in the dark, but did not grow under blue light.
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[[File:K24 1.png|600px|thumb|center|Figure1. <b>(1) pDawn(cI)-X174E-T1</b>; (2) pDawn(cI-LVA)-X174E-LVA-T1; (3) pDawn(cI)-LKD-T1; (4) pDawn(cI-LVA)-RBS070-LKD-LVA-T1 were constructed successfully.]]<p></p> [[File:K24 2.png|600px|thumb|center|Figure2. Growth condition of <i>E.coli </i> TOP10-pDawn(cI)-X174E-T1 in the dark and under blue light.]]
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<p></p> As shown in Figure 1, we successfully constructed pDawn(cI)-X174E-T1 and transformed it into <i> E. coli</i> TOP10, selecting bacteria colonies and culturing in different growth conditions. As shown in Figure 2, the 9<sup>th</sup> colony of <i>E.coli</i> TOP10-pDawn(cI)-X174E-T1 grew in the dark, but did not grow under blue light.
 
<h3>References</h3>
 
<h3>References</h3>
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<p>Robert Ohlendorf 1 , Roee R. Vidavski 2 , Avigdor Eldar 2, Keith Moffat 3,4 and Andreas Möglich 1, 3.From Dusk till Dawn: One-Plasmid Systems for Light-Regulated Gene Expression.
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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.
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</p >
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<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 >
 
</p >

Latest revision as of 14:36, 12 October 2022

pDawn(cl)-X174E-T1

Description

This composite part is a generator consisting of pDawn(cI) (BBa_K4325010), X174E (BBa_K1835500) and T1 terminator (BBa_K3033016).

Usage

We inserted the blue light responsive system pDawn(cI) (BBa_K4325010) and the lysis gene X174E (BBa_K1835500) into the pSEVA331 expression vector, which was introduced into E. coli TOP10. The bacterial colonies that grew in the dark but did not grow under blue light were screened to verify the responsiveness of pDawn(cI) to 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(cl)-X174E-T1 by evaluating the response to blue light.

Figure1. (1) pDawn(cI)-X174E-T1; (2) pDawn(cI-LVA)-X174E-LVA-T1; (3) pDawn(cI)-LKD-T1; (4) pDawn(cI-LVA)-RBS070-LKD-LVA-T1 were constructed successfully.

Figure2. Growth condition of E.coli TOP10-pDawn(cI)-X174E-T1 in the dark and under blue light.

As shown in Figure 1, we successfully constructed pDawn(cI)-X174E-T1 and transformed it into E. coli TOP10, selecting bacteria colonies and culturing in different growth conditions. As shown in Figure 2, the 9th colony of E.coli TOP10-pDawn(cI)-X174E-T1 grew in the dark, but did not grow under blue light.

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).