Difference between revisions of "Part:BBa K5115053"
 
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<partinfo>BBa_K5115053 short</partinfo>
 
<partinfo>BBa_K5115053 short</partinfo>
  
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<html><img style="float:right;width:128px" src="https://static.igem.wiki/teams/5115/czh/mineral-logo.svg" alt="contributed by Fudan iGEM 2023"></html>
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<html><img style="float:right;width:128px" src="https://static.igem.wiki/teams/5115/czh/mineral-logo.svg" alt="contributed by Fudan iGEM 2024"></html>
 
__TOC__
 
__TOC__
 
===Introduction===
 
===Introduction===
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This composite part combines all the subunit, except hoxU, of the hydrogenase in our ribozyme-assisted polycistronic co-expression system：pRAP. To learn more about the hydrogenase, please check [https://parts.igem.org/Part:BBa_K5115020 BBa_K5115020(hox and hyp operon)]. To get more information about pRAP, please check [https://2022.igem.wiki/fudan/parts part wiki of 2022 Fudan iGEM ].
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This composite part combines all the subunits, except hoxU, of the hydrogenase in our ribozyme-assisted polycistronic co-expression system：pRAP. Actually, compared with [https://parts.igem.org/Part:BBa_K5115052 ribozyme connected hox and hyp, without hoxF], there is only one difference: one deleted hoxF and another deleted hoxU. The deleted subunits are all used to link with EP. EP ligation of different subunits leads to different effects of hydrogenase's entry into the carboxysome, which further affects the effect of our overall design. After comparison between experiments of these two designs, we make sure that the hoxF-linking-EP one is more satisfying. So, to learn more about the final adopted part, please click on [https://parts.igem.org/Part:BBa_K5115052 ribozyme connected hox and hyp, without hoxF].
  
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===Characterization===
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====Agarose gel electrophoresis====
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{|
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| <html><img style="width:400px" src="https://static.igem.wiki/teams/5115/registry/ribozyme-connected-hox-and-hyp-without-hoxu.png" alt="contributed by Fudan iGEM 2024"></html>
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|-
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| '''Figure 1. Agarose gel electrophoresis of PCR products amplified from one ''E. coli'' (DH5α) colony.   
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M: DNA Marker. Lanes 1,3-8: Amplification of specific regions corresponding to hoxF, hoxY, hoxH, hoxW, hoxI, hypA, and hypB, demonstrating successful assembly and integrity of the ribozyme-connected hox and hyp, without hoxU (no band in lane 2) as designed. Primers for these PCR are listed on https://2024.igem.wiki/fudan/parts.
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'''
  
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|}
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===Sequence and Features===
 
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<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>

Latest revision as of 12:57, 2 October 2024


ribozyme connected hox and hyp, without hoxU

contributed by Fudan iGEM 2024

Introduction

This composite part combines all the subunits, except hoxU, of the hydrogenase in our ribozyme-assisted polycistronic co-expression system：pRAP. Actually, compared with ribozyme connected hox and hyp, without hoxF, there is only one difference: one deleted hoxF and another deleted hoxU. The deleted subunits are all used to link with EP. EP ligation of different subunits leads to different effects of hydrogenase's entry into the carboxysome, which further affects the effect of our overall design. After comparison between experiments of these two designs, we make sure that the hoxF-linking-EP one is more satisfying. So, to learn more about the final adopted part, please click on ribozyme connected hox and hyp, without hoxF.

Characterization

Agarose gel electrophoresis

contributed by Fudan iGEM 2024
Figure 1. Agarose gel electrophoresis of PCR products amplified from one E. coli (DH5α) colony.

M: DNA Marker. Lanes 1,3-8: Amplification of specific regions corresponding to hoxF, hoxY, hoxH, hoxW, hoxI, hypA, and hypB, demonstrating successful assembly and integrity of the ribozyme-connected hox and hyp, without hoxU (no band in lane 2) as designed. Primers for these PCR are listed on https://2024.igem.wiki/fudan/parts.

Sequence and Features

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 654
    Illegal BglII site found at 1472
    Illegal BglII site found at 1765
    Illegal BamHI site found at 2059
    Illegal XhoI site found at 99
    Illegal XhoI site found at 2266
    Illegal XhoI site found at 4505
    Illegal XhoI site found at 6334
    Illegal XhoI site found at 7496
    Illegal XhoI site found at 9335
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 533
    Illegal NgoMIV site found at 1071
    Illegal NgoMIV site found at 1281
    Illegal NgoMIV site found at 1593
    Illegal NgoMIV site found at 2359
    Illegal NgoMIV site found at 3039
    Illegal NgoMIV site found at 5646
    Illegal AgeI site found at 6821
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 4159
    Illegal BsaI.rc site found at 702
    Illegal BsaI.rc site found at 1206


References