Difference between revisions of "Part:BBa K3050000"
 
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<partinfo>BBa_K3050000 short</partinfo>
 
<partinfo>BBa_K3050000 short</partinfo>
  
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Magainin-2 expressed inside E.coli combines to cell membrane and damages it.In our project, we aimed to make BL21(DE3) lyse easier by transforming BBa_K3050000 on pSB1C3.
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Magainin-2 expressed inside E.coli combines to cell membrane and damages it. In our project, we aimed to make BL21(DE3) lyse easier by transforming BBa_K3050000 on pSB1C3.
  
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<!-- Add more about the biology of this part here
 
 
===Usage and Biology===
 
===Usage and Biology===
  
 
This is a composite part that includes T7 promotor + RBS + mag-2 + GFP + double terminators.
 
This is a composite part that includes T7 promotor + RBS + mag-2 + GFP + double terminators.
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You can use this part when you want to weaken cell membrane of E.coli,also you can visually know when it expressed.
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As the magainin-2 and GFP is under T7 promotor, you can easily induce it by adding IPTG.
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We recommend 300 μM IPTG for directly putting on colonies on plates, and 500 μM for culture solution.
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You want to know more what is it like when IPTG is added?
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Read this page more or see also: https://parts.igem.org/Part:BBa_I746909:Experience
  
 
BBa_I746909 is linerized by Inverse PCR to be combined with magainin-2 sequense by In-Fusion reaction.
 
BBa_I746909 is linerized by Inverse PCR to be combined with magainin-2 sequense by In-Fusion reaction.
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GFP part is on downstream of T7 promotor and magainin-2, hence we're quite sure that this result indicates that magainin-2 is expressed at the same time.  
 
GFP part is on downstream of T7 promotor and magainin-2, hence we're quite sure that this result indicates that magainin-2 is expressed at the same time.  
  
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<div class="students">
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[[File:T--Gunma--BBa_K3050000.png|600px|center|]]
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    <div class="card">
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Fig.1 BL21(DE3) pSB1C3 BBa_K3050000 expressed under visible light with negative control without IPTG induction
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        <img src="https://2019.igem.org/wiki/images/b/b9/T--Gunma--BBa_K3050000.png">
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    </div>
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</div>Fig.1 BL21(DE3) pSB1C3 BBa_K3050000 expressed under visible light with negative control without IPTG induction
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[[File:T--Gunma--BBa_K3050000_2.png|600px|center|]]
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Fig.2 pSB1C3 BBa_K3050000 expressed under UV light with negative control without IPTG induction
  
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<div class="clear"></div>
 
  
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<div class="students">
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[[File:T--Gunma--BBa_K3050000_3.png|600px|center|]]
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    <div class="card">
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Fig.3 Four biological replicates all highly expressed
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        <img src="https://2019.igem.org/wiki/images/e/e2/T--Gunma--BBa_K3050000_2.png">
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    </div>
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</div>Fig.2 pSB1C3 BBa_K3050000 expressed under UV light with negative control without IPTG induction
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<div class="clear"></div>
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[[File:T--Gunma--K3050000 graph.PNG|600px|center|]]
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Fig.4 500 μM IPTG induced BBa K3050000, strongly emitting green fluorescent light
  
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<div class="students">
 
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    <div class="card">
 
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        <img src="https://2019.igem.org/wiki/images/0/05/T--Gunma--BBa_K3050000_3.png">
 
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    </div>
 
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</div>Fig.3 Four biological replicates all highly expressed
 
  
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<div class="clear"></div>
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<!-- Add more about the biology of this part here
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===Usage and Biology===
  
 
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<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
 
<partinfo>BBa_K3050000 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K3050000 SequenceAndFeatures</partinfo>

Latest revision as of 23:28, 21 October 2019


pT7+RBS+mag-2+GFP+double_terminator

Magainin-2 expressed inside E.coli combines to cell membrane and damages it. In our project, we aimed to make BL21(DE3) lyse easier by transforming BBa_K3050000 on pSB1C3.

Usage and Biology

This is a composite part that includes T7 promotor + RBS + mag-2 + GFP + double terminators. You can use this part when you want to weaken cell membrane of E.coli,also you can visually know when it expressed. As the magainin-2 and GFP is under T7 promotor, you can easily induce it by adding IPTG. We recommend 300 μM IPTG for directly putting on colonies on plates, and 500 μM for culture solution. You want to know more what is it like when IPTG is added? Read this page more or see also: https://parts.igem.org/Part:BBa_I746909:Experience

BBa_I746909 is linerized by Inverse PCR to be combined with magainin-2 sequense by In-Fusion reaction. https://parts.igem.org/Part:BBa_I746909

BL21(DE3) transformed pSB1C3 BBa_K3050000 successfully grew and emit strong green fluorescent light as shown in pictures. Thus we succeeded to prove Inverse PCR,In-Fusion reaction,and transformation went great.

GFP part is on downstream of T7 promotor and magainin-2, hence we're quite sure that this result indicates that magainin-2 is expressed at the same time.

T--Gunma--BBa K3050000.png

Fig.1 BL21(DE3) pSB1C3 BBa_K3050000 expressed under visible light with negative control without IPTG induction


T--Gunma--BBa K3050000 2.png

Fig.2 pSB1C3 BBa_K3050000 expressed under UV light with negative control without IPTG induction


T--Gunma--BBa K3050000 3.png

Fig.3 Four biological replicates all highly expressed


T--Gunma--K3050000 graph.PNG

Fig.4 500 μM IPTG induced BBa K3050000, strongly emitting green fluorescent light


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 190