| Types | DnaRegion
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| Roles | plasmid_vector
Plasmid_Backbone
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Description
This plasmid was derived from pSac-Cm by insertion of biobrick compatible restriction sites (prefixes and suffixes), a terminator (BBa_B0015) after the suffixes sequences, and red fluorescent protein sequence (RFP) in between the prefix and suffix in its multiple cloning sites (MCS). New biobricks can be inserted into this vector by replacement of the RFP biobrick, and selection of the white colonies. This backbone has a multi host replication origin and replicates in E. coli. The plasmid is designed to integrate a cloned insert into the B. subtilis chromosome via double recombination between plasmid and chromosomal sacA sequences, and in order to check for integration a simple Phenol Red Sucrose test can be perform, if sacA is no longer functional, they will remain ph7, if they are still able to digest sucrose their pH will be under 5.
Notes
We found that that the integration plasmid that groningen send to the registry were unable to integrate in B. subtilis. This is due to the high number of mutations in the sacA integration region.
Source
Groningen iGEM 2012 team
