pSB2K3 is a variable copy number plasmid carrying kanamycin resistance. The plasmid contains the F' replication origin and also the P1 lytic replication origin. Genes encoding proteins that are required to activate the P1 lytic replication origin are carried on the plasmid under the control of a lacI-regulated promoter. When grown in cells producing LacI the plasmid is maintained at very low copy (<10 per cell). When induced with IPTG the P1 lytic replication proteins are produced and the P1 lytic replication origin takes over, raising the copy number to very high levels (>100++ per cell). The copy control features of pSB2K3 are useful for cloning "difficult" parts while still being able to prepare large amounts of DNA. Additionally, pSB2K3 has terminators bracketing its MCS which are designed to prevent transcription from *inside* the MCS from reading out into the vector. The efficiency of these terminators is known to be < 100%. Ideally we would construct a future set of terminators for bracketing a MCS that were 100% efficient in terminating both into and out of the MCS region.
*** pSCANS via BNL *** The pSB2 series plasmids are based on the pSCANS-5 plasmid from John Dunn's group at Brookhaven National Laboratory. The pSCANS plasmids are introduced here.
MIT SBWG modification of pSCANS-5 from BNL
pSB2K3-derived BioBrick plasmid free of many restriction sites
Derived from pSB2K3 with many restriction enzyme sites removed by site-directed mutagenesis (~2 dozen mutations). Plasmid is useful when there is a need to cut with a variety of different restriction enzymes. Most symmetric cutters have been removed, and some non-symmetric cutters have also including AarI, AcuI, BbsI, BbvCI, BciVI, BsaI, BsmBI, EarI, N.BstNBI, and SgrAI. Works in exactly same way as pSB2K3. See <bb_part>pSB2K3</bb_part> for more information. Note that the VF2 site has a single base mutation in the middle of it. However, VF2 has been used successfully as a primer for PCR with this plasmid.