Difference between revisions of "Part:BBa K3332005"

 
 
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<partinfo>BBa_K3332005 short</partinfo>
 
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iNAP is fused at C-terminal with BrkA anchoring protein. We use K880005 to construct the expression system and anchor iNAP on the surface of E.coli.
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iNAP is fused at C-terminal with BrkA anchoring protein. We use <partinfo>BBa_K880005</partinfo> to construct the expression system and anchor iNAP on the surface of ''E.coli''.
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===Biology===
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&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;BrkA is an anchor protein from ''Bordetella pertussis'' having β-barrel structure. It can anchor its passenger protein to the cell membrane and has been widely used in cell-surface display. iNap is a chimera of circularly permuted YFP (cpYFP) and the NADP(H) binding domain of Rex from Thermus aquaticus (T-Rex). Upon NADPH binding, iNap shows apparent fluorescence changes. iNap is fused at C terminal with BrkA so that iNap can be displayed on the surface of ''E. coli''.<ref>Zou Y, Wang A, Shi M, et al. Analysis of redox landscapes and dynamics in living cells and in vivo using genetically encoded fluorescent sensors[J]. Nat Protoc, 2018, 13(10): 2362-2386.</ref><ref>http://2016.igem.org/Team:TJUSLS_China</ref>
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    <figure>
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        <img src="https://2020.igem.org/wiki/images/9/94/T--XMU-China--XMU-China_2020-iNAP_Mechanism.png" width="50%" style="float:center">
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        <figcaption>
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        <p style="font-size:1rem">
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        </p>
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        </figcaption>
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    </figure>
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:'''Fig 1.''' Mechanism of iNap on the surface of ''E. coli''
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===Usage===
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&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;Here, we used <partinfo>BBa_K880005</partinfo> to construct the expression system and obtained the composite part <partinfo>BBa_K3332050</partinfo>, which may achieve surface display of iNap on our engineering bacteria.  Due to the limited time, we did not get the gene in time. As a result, there is a lack of data about this part. The progress of this part remains in the stage of design.
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        <img src="https://2020.igem.org/wiki/images/c/ca/T--XMU-China--XMU-China_2020-J23100_B0034_inap-brkA_B0015.png" width="50%" style="float:center">
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        <figcaption>
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        <p style="font-size:1rem">
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        </figcaption>
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:'''Fig 2.''' Gene circuit of iNap-BrkA
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===References===
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<references/>
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===Usage and Biology===
 
  
 
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Latest revision as of 22:51, 27 October 2020


iNAP-BrkA

iNAP is fused at C-terminal with BrkA anchoring protein. We use BBa_K880005 to construct the expression system and anchor iNAP on the surface of E.coli.


Biology

        BrkA is an anchor protein from Bordetella pertussis having β-barrel structure. It can anchor its passenger protein to the cell membrane and has been widely used in cell-surface display. iNap is a chimera of circularly permuted YFP (cpYFP) and the NADP(H) binding domain of Rex from Thermus aquaticus (T-Rex). Upon NADPH binding, iNap shows apparent fluorescence changes. iNap is fused at C terminal with BrkA so that iNap can be displayed on the surface of E. coli.[1][2]

Fig 1. Mechanism of iNap on the surface of E. coli


Usage

        Here, we used BBa_K880005 to construct the expression system and obtained the composite part BBa_K3332050, which may achieve surface display of iNap on our engineering bacteria. Due to the limited time, we did not get the gene in time. As a result, there is a lack of data about this part. The progress of this part remains in the stage of design.

Fig 2. Gene circuit of iNap-BrkA


References

  1. ↑ Zou Y, Wang A, Shi M, et al. Analysis of redox landscapes and dynamics in living cells and in vivo using genetically encoded fluorescent sensors[J]. Nat Protoc, 2018, 13(10): 2362-2386.
  2. ↑ http://2016.igem.org/Team:TJUSLS_China



Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 272
    Illegal XhoI site found at 298
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 38
    Illegal NgoMIV site found at 1459
    Illegal NgoMIV site found at 1888
    Illegal NgoMIV site found at 2524
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 2492