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Plasmid

Part:BBa_I52001:Design

Designed by: Reshma Shetty   Group:   (2007-01-29)

ccdB and nonfunctional pUC19 derived high copy origin


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 435
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 323

Design Notes

The BioBrick standard vectors are designed to be easy to use for their most common purpose: assembly of BioBrick standard biological parts. To meet this requirement, we included BBa_P1016 within the BioBrick cloning site. BBa_P1016 encodes the positive selection marker ccdB. Positive selection markers prevent one of the most common problems during assembly of BioBrick parts: contamination of the ligation reaction with uncut plasmid DNASambrook-2001. Any cells transformed with the uncut plasmid DNA produce the lethal protein ccdB and dieBernard-Gene-1994 Bernard-Gene-1995 Bernard-Biotechniques-1996. Note, however, that the drawback of this solution is that inclusion of this part requires that users propagate both the base vector and any derived vectors in E. coli strains tolerant of ccdB expression such as DB3.1Bernard-J-Mol-Biol-1992 Miki-J-Mol-Biol-1992.

Source

BBa_P1016 (ccdB positive selection marker) and BBa_I50020 (nonfunctional high copy origin derived from pUC19).

References

  • Bernard P, Gabant P, Bahassi EM, Couturier M, Université Libre de Bruxelles. Positive-selection vectors using the F plasmid ccdB killer gene." Gene 1994 Oct 11;148(1):71-4. pmid:7926841. [http://www.ncbi.nlm.nih.gov/pubmed/7926841 Pubmed]
  • Bernard P, Gabant P, Université Libre de Bruxelles. Cloning and/or sequencing vector US Patent Number 5,910,438, 1999. [http://www.google.com/patents?vid=USPAT5910438 Google Patents]
  • Bernard P, Gabant P, Université Libre de Bruxelles. Cloning and/or sequencing vector US Patent Number 6,180,407 B1, 2001. [http://www.google.com/patents?vid=USPAT6180407 Google Patents]
  • Bernard P, Gabant P, Université Libre de Bruxelles. Cloning and/or sequencing vector US Patent Number 7,176,029 B2, 2007. [http://www.google.com/patents?vid=USPAT7176029 Google Patents]
  • Bahassi, et al., Université Libre de Bruxelles. Interactions of CcdB with DNA gyrase. Inactivation of Gyra, poisoning of the gyrase-DNA complex, and the antidote action of CcdA., J. Biol. Chem. 1999 274: 10936-10944. pmid:10196173. [http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=10196173&query_hl=2&itool=pubmed_ExternalLink Pubmed]

<biblio>

  1. Bernard-J-Mol-Biol-1992 pmid=1324324
  2. Miki-J-Mol-Biol-1992 pmid=1316444
  3. Bernard-Gene-1995 pmid=7557407
  4. Bernard-Biotechniques-1996 pmid=8862819

</biblio>