Difference between revisions of "Part:BBa K525720:Design"

(→‎Design Notes)
(→‎Design Notes)
 
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*This is the complete sequence of NAD<sup>+</sup>  -dependent DNA ligase.
 
*This is the complete sequence of NAD<sup>+</sup>  -dependent DNA ligase.
 
*It is designed in [https://parts.igem.org/Assembly_standard_25 RFC 25]
 
*It is designed in [https://parts.igem.org/Assembly_standard_25 RFC 25]
 +
**NgoMIV restriction site downstream the start codon to easily fuse an N-terminal domain to the coding sequence
 +
**AgeI restriction site upstream of stop codon to easily fuse a C-terminal domain to the coding sequence
  
 
===Source===
 
===Source===

Latest revision as of 21:57, 21 September 2011

DNA ligase from Escherichia coli (LigA)


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 1422
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 4
    Illegal AgeI site found at 2020
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 1636


Design Notes

  • This is the complete sequence of NAD+ -dependent DNA ligase.
  • It is designed in RFC 25
    • NgoMIV restriction site downstream the start codon to easily fuse an N-terminal domain to the coding sequence
    • AgeI restriction site upstream of stop codon to easily fuse a C-terminal domain to the coding sequence

Source

  • PCR on the Escherichia coli TOP 10 DNA cloned into BioBrick Vectors
  • Primers with BioBrick Freiburg prefix in the fwd primer. Freiburg Suffix in the rev primer.
    • fwd: 5'-acgtgaattcgcggccgcttctagatggccggcgaatcaatcgaacaacaactgac-3'
    • rev: 5'-acgtctgcagcggccgctactagtattaaccggtgctacccagcaaacgc-3'

References