Synthetic Biology:Vectors/Single copy plasmid

See Vectors for information on vector nomenclature, existing vectors and vectors that we would like constructed.

Construction of single copy BioBricks vector

See notes on bacterial artificial chromosomes.

To do list

• One of the things need for this project is BioBricked antibiotic resistance cassettes. This is also a prerequisite for the standard strain project. Tom has ordered primers and is planning on cloning several resistance cassettes.
• Reshma: specify F plasmid backbone with no internal BioBricks restriction sites based on the part of pBAC108L used in pBACe3.6. Ask Randy to get it synthesized?
• ccdB is already in BioBricks form
• Plan assembly of vector modules.

To be decided

• If all the vector components are specified in BioBricks format, how do we construct a BioBricks insertion site?
  • Blunt-end ligation?
  • Other restriction enzyme sites?
  • PCR
• What is the best way to assemble a pUC backbone with the F plasmid backbone such that the pUC backbone is flanked by BioBricks sites?
  • PCR
  • Use special sites for vector construction

To be determined

• Are we sure that F plasmids are really at 1-2 copies per cell? Why was pSB2K3-1 measured to be higher than that?
• What parts of the F plasmid are responsible for integration onto the genome? Can we omit them?
• Many of the existing BACs only seem to have a partial sopC CDS, do we want the rest?