Part:BBa_I50022:Design

Minimal pUC19-derived high copy replication origin

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 19
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Design Notes

BioBrick standard vectors are also designed to be easy to purify. To meet this requirement, we included a pUC19-derived origin (BBa_I50022) in addition to the ccdB selection marker within the BioBrick cloning site of BioBrick standard vectorsVieira-Gene-1982 Norrander-Gene-1983 Yanisch-Perron-Gene-1985. The high copy origin encoded by BBa_I50022 means that both base vector DNA and any derived vector DNA are easily purified in large quantities, irrespective of whether the vector replication origin is low copy or notCabello-Nature-1976 Ioannou-Nat-Genet-1994. Cloning a BioBrick part into the BioBrick cloning site removes the high copy origin in the cloning site thereby restoring replication control to the vector origin.

BBa_I50022 was originally designed as BBa_I50020. However, several introduced mutations to the pUC19 origin sequence rendered BBa_I50020 nonfunctional as a replication origin. Hence, these introduced mutations were reverted to make BBa_I50022, a functional high copy replication origin.

Source

pSB1A3 and BBa_I50020 served as the basis of the design of BBa_I50022.

References

<biblio>

1. Vieira-Gene-1982 pmid=6295879
2. Norrander-Gene-1983 pmid=6323249
3. Yanisch-Perron-Gene-1985 pmid=2985470
4. Cabello-Nature-1976 pmid=765836
5. Ioannou-Nat-Genet-1994 pmid=8136839

</biblio>