Difference between revisions of "Part:BBa K4011018"

 
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pTEF1-SaACS2-tADH1-pRPL8B-AeAT9-tSSA1 is an expression cassette in <i>S. cerevisiae</i> capable of expressing AeAT9 and SaACS2, which will synthesize ethyl-acetate from acetate and ethanol. SaACS2 will convert acetate into acetyl-CoA, and AeAT9 will convert acetyl-CoA and ethanol to ethyl-acetate. pTEF1-SaACS2-tADH1 <partinfo>BBa_K4011016</partinfo> and pRPL8B-AeAT9-tSSA1 <partinfo>BBa_K4011017</partinfo> were used to construct pTEF1-SaACS2-tADH1-pRPL8B-AeAT9-tSSA1.
 
pTEF1-SaACS2-tADH1-pRPL8B-AeAT9-tSSA1 is an expression cassette in <i>S. cerevisiae</i> capable of expressing AeAT9 and SaACS2, which will synthesize ethyl-acetate from acetate and ethanol. SaACS2 will convert acetate into acetyl-CoA, and AeAT9 will convert acetyl-CoA and ethanol to ethyl-acetate. pTEF1-SaACS2-tADH1 <partinfo>BBa_K4011016</partinfo> and pRPL8B-AeAT9-tSSA1 <partinfo>BBa_K4011017</partinfo> were used to construct pTEF1-SaACS2-tADH1-pRPL8B-AeAT9-tSSA1.
  
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==Usage and Biology==
  
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SaACS2 is an acetyl-CoA synthase from <i>Salmonella enterica</i>. Its natural function is to convert acetate into acetyl-CoA under anaerobic conditions. Its enzymatic rate is around 50 times that of ACS1 found in <i>S. cerevisiae</i>. AeAT9 is an alcohol acyltransferase from <i>Actinidia eriantha</i>, otherwise known as kiwifruit. Its general function is to convert alcohols into esters. The fruity smell one smells from kiwifruit it largely a result of AeAT9.  pTEF1 is a relatively strong promoter, tADH1 is a medium strength terminator, pRPL8B is a medium strength promoter, and tADH1 is a relatively strong terminator all characterized by Lee et al in 2021.
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===Usage and Biology===
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===Source===
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pTEF1, tADH1, pRPL8B, and tSSA1 all come from <i>S. cerevisiae</i>, while SaACS2 comes from <i>Salmonella enterica</i> and AeAT9 comes from <i>Actinidia eriantha</i>.
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==Design Considerations==
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1. All codons were optimized for <i>S. cerevisiae</i> based on <i>S. cerevisiae</i> codon bias.
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2. Homologous recombination of <i>S. cerevisiae</i> was avoided by choosing different promoters and terminators for each expression casette (SaACS2 and AeAT9).
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==Characterization==
  
 
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Revision as of 12:58, 21 October 2021


pTEF1-SaACS2-tADH1-pRPL8B-AeAT9-tSSA1

pTEF1-SaACS2-tADH1-pRPL8B-AeAT9-tSSA1 is an expression cassette in S. cerevisiae capable of expressing AeAT9 and SaACS2, which will synthesize ethyl-acetate from acetate and ethanol. SaACS2 will convert acetate into acetyl-CoA, and AeAT9 will convert acetyl-CoA and ethanol to ethyl-acetate. pTEF1-SaACS2-tADH1 BBa_K4011016 and pRPL8B-AeAT9-tSSA1 BBa_K4011017 were used to construct pTEF1-SaACS2-tADH1-pRPL8B-AeAT9-tSSA1.

Usage and Biology

SaACS2 is an acetyl-CoA synthase from Salmonella enterica. Its natural function is to convert acetate into acetyl-CoA under anaerobic conditions. Its enzymatic rate is around 50 times that of ACS1 found in S. cerevisiae. AeAT9 is an alcohol acyltransferase from Actinidia eriantha, otherwise known as kiwifruit. Its general function is to convert alcohols into esters. The fruity smell one smells from kiwifruit it largely a result of AeAT9. pTEF1 is a relatively strong promoter, tADH1 is a medium strength terminator, pRPL8B is a medium strength promoter, and tADH1 is a relatively strong terminator all characterized by Lee et al in 2021.

Source

pTEF1, tADH1, pRPL8B, and tSSA1 all come from S. cerevisiae, while SaACS2 comes from Salmonella enterica and AeAT9 comes from Actinidia eriantha.

Design Considerations

1. All codons were optimized for S. cerevisiae based on S. cerevisiae codon bias.

2. Homologous recombination of S. cerevisiae was avoided by choosing different promoters and terminators for each expression casette (SaACS2 and AeAT9).

Characterization

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 3375
    Illegal BglII site found at 3582
    Illegal BglII site found at 4200
    Illegal XhoI site found at 2426
    Illegal XhoI site found at 4699
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 3609
    Illegal AgeI site found at 1987
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 2412
    Illegal BsaI site found at 4685
    Illegal BsaI.rc site found at 205
    Illegal BsaI.rc site found at 2662
    Illegal BsaI.rc site found at 4935
    Illegal SapI.rc site found at 4447