Difference between revisions of "Part:BBa K1716000:Design"

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===Design Notes===
 
===Design Notes===
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The coding sequence for ''E. coli'' phage (GeneBank: KT232076.1) derived lambda recombinase was optimized for expression in Bacillus subtilis using JCat.
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pMiniMAD contains an internal BsaI cut site within the ampicillin resistance gene – this can be avoided when using BsaI by ensuring minimal homology of the internal 4 bp overhang with other designed overhangs and performing an additional final ligation step following Golden Gate assembly.
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Alternatively, the BsaI recognition sequence could be removed by point mutation to a synonymous codon, by site-directed mutagenesis or using an additional internal set of primers.
  
 
===Source===
 
===Source===

Revision as of 14:52, 8 October 2023


Lambda Beta recombinase optimized for expression in B. subtilis


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 245


Design Notes

pMiniMAD contains an internal BsaI cut site within the ampicillin resistance gene – this can be avoided when using BsaI by ensuring minimal homology of the internal 4 bp overhang with other designed overhangs and performing an additional final ligation step following Golden Gate assembly.

Alternatively, the BsaI recognition sequence could be removed by point mutation to a synonymous codon, by site-directed mutagenesis or using an additional internal set of primers.

Source

Gene product from E. coli phage lambda.

References

H. H. Wang, et al., “Programming Cells by Multiplex Genome Engineering and Accelerated Evolution,” Nature, Vol. 460, No. 7257, 2009, pp. 894-898. doi:10.1038/nature08187