Difference between revisions of "Part:BBa K3784002"

 
 
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<partinfo>BBa_K3784002 short</partinfo>
 
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fldC is from the fld gene cluster of Clostridium sporogenes, involved in the conversion of tryptophan to 3-Indolepropionic acid, which encodes acyl-CoA dehydrogenase.
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The fldC is from the FLD gene cluster of Clostridium sporogenes, involved in the conversion of tryptophan to 3-Indolepropionic acid. In the five-step metabolic pathway we reconstructed, fldC encodes subunit of 2-hydroxyacyl-CoA dehydratase family protein, combined with another subunit encoded by fldB to become 2-hydroxyacyl-CoA dehydratase family protein, which catalyze indole lactic acid to produce indole acrylic acid.
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===Characterization===
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We successfully inserted the fldC gene into the His-6p-MBP-RSFD plasmid and transformed it into E. coli BL21(DE3) and induced it to express. After extracting total RNA, we obtained its cDNA by reverse transcription. We used cDNA as a template to amplify the fragment, then checked it by agarose gel electrophoresis. We chose rsmA (16S rRNA m(6)2A1518, m(6)2A1519 dimethyltransferase) as the internal reference gene and set a negative control. The fldC is well expressed and the expression levels of internal reference genes in the experimental group (rsmAE) and the control group (rsmAC) were consistent.
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[[File:BNUZ-fldC.png]]
  
 
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Latest revision as of 01:58, 20 October 2021


fldC (Encoding 2-hydroxyacyl-CoA dehydratase family protein)

The fldC is from the FLD gene cluster of Clostridium sporogenes, involved in the conversion of tryptophan to 3-Indolepropionic acid. In the five-step metabolic pathway we reconstructed, fldC encodes subunit of 2-hydroxyacyl-CoA dehydratase family protein, combined with another subunit encoded by fldB to become 2-hydroxyacyl-CoA dehydratase family protein, which catalyze indole lactic acid to produce indole acrylic acid.

Characterization

We successfully inserted the fldC gene into the His-6p-MBP-RSFD plasmid and transformed it into E. coli BL21(DE3) and induced it to express. After extracting total RNA, we obtained its cDNA by reverse transcription. We used cDNA as a template to amplify the fragment, then checked it by agarose gel electrophoresis. We chose rsmA (16S rRNA m(6)2A1518, m(6)2A1519 dimethyltransferase) as the internal reference gene and set a negative control. The fldC is well expressed and the expression levels of internal reference genes in the experimental group (rsmAE) and the control group (rsmAC) were consistent.

BNUZ-fldC.png

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal XbaI site found at 799
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal XbaI site found at 799
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal XbaI site found at 799
  • 1000
    COMPATIBLE WITH RFC[1000]