Difference between revisions of "Part:BBa K3268002"
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4. Any inserted protein-coding genes not containing HindII and BamHI sites in their coding sequences can then be amplified and cloned into our composite expression module improved from BBa_J364007.
 
4. Any inserted protein-coding genes not containing HindII and BamHI sites in their coding sequences can then be amplified and cloned into our composite expression module improved from BBa_J364007.
 
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Revision as of 15:42, 20 October 2019


Strong expression of eforRed in E. coli


To make a composite module that can highly express any inserted protein-coding genes, we had designed and constructed this strongly expressed eforRed in E. coli as a proof of our concept.
1. The eforRed coding region was PCR amplified from BBa_K592012 using high-fidelity enzyme KOD-Plus.
2. The strong promoter along with BBa_B0034 RBS and BBa_B0015 transcriptional terminator on pSB1C3 plasmid backbone were PCR amplified from BBa_J364007 using high-fidelity enzyme KOD-Plus.
3. In order to make the strong promoter (along with BBa_B0034 RBS and BBa_B0015 transcriptional terminator on pSB1C3 plasmid backbone) useful to highly express inserted protein-coding genes, we had designed two PCR primers with HindII and BamHI sites respectively to generate a composite expression module without the original GFP coding sequence.
4. Any inserted protein-coding genes not containing HindII and BamHI sites in their coding sequences can then be amplified and cloned into our composite expression module improved from BBa_J364007.


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal prefix found in sequence at 2047
    Illegal PstI site found at 14
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 2047
    Illegal NheI site found at 2075
    Illegal NheI site found at 2098
    Illegal PstI site found at 14
    Illegal NotI site found at 7
    Illegal NotI site found at 2053
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 2047
    Illegal BamHI site found at 2124
    Illegal XhoI site found at 1031
    Illegal XhoI site found at 1923
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal prefix found in sequence at 2047
    Illegal PstI site found at 14
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal prefix found in sequence at 2047
    Illegal XbaI site found at 2062
    Illegal PstI site found at 14
  • 1000
    COMPATIBLE WITH RFC[1000]