pMC_0_7+8_panS_specResLVL1
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal XbaI site found at 1199
Illegal PstI site found at 3734 - 12INCOMPATIBLE WITH RFC[12]Illegal PstI site found at 3734
Illegal NotI site found at 1
Illegal NotI site found at 5738 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 4897
Illegal XhoI site found at 1249 - 23INCOMPATIBLE WITH RFC[23]Illegal XbaI site found at 1199
Illegal PstI site found at 3734 - 25INCOMPATIBLE WITH RFC[25]Illegal XbaI site found at 1199
Illegal PstI site found at 3734
Illegal NgoMIV site found at 48
Illegal NgoMIV site found at 190 - 1000COMPATIBLE WITH RFC[1000]
Overview
This part is contained in the green expansion, a range of parts from iGEM Marburg 2020 that enables user of the Marburg Collection 2.0 to design MoClo compatible vectors for cyanobacteria as well as to engineer the genome of several cyanobacterial species.
The green expansion
The green expansion in the Marburg Collection 2.0 features the world's first MoClo compatible shuttle vector for cyanobacteria (BBa_K3228069). This composite part contains the origins of replication ColE1 (for cloning) and the ori of panS (for maintenance in cyanobacteria) as well as a spectinomycin cassette. It can be used in the Marburg Collection like any other composite part of the type 7+8 (antibiotic cassette + origin of replication).
This expansion also contains all the parts needed for the genomic integration of one or multiple genes in cyanobacteria. It convinces with a striking flexibility and a very intuitive workflow for the de novo assembly of your plasmid of choice. It encompasses 5 different neutral integration sites to choose from: three conventional sites frequently used in the cyanobacterial community (NSI to NSIII) as well as our own rationally designed artificial neutral integration site options (a.N.S.o. 1 and 2). These sites show no transcriptional activity from neighboring regions and are therefore completely orthogonal. Additionally we offer 4 different antibiotic markers to use (chloramphenicol, gentamycin, spectinomycin and kanamycin). In theory with this setup up to 20 genes could be introduced into a cyanobacterial strain.
Thanks to the flexible design this expansion can also be used for the genomic modification of any chassis after the introduction of new species specific LVL 0 integration sites to our Marburg Collection 2.0. As the workflow to build new homologies is a bit more intricate compared to the one pot on step assembly of our other parts due to the internal BsmbI cutting site, we described the workflow for that in our design section (See Design: Design of the A.N.S.O.s).
The green expansion proves a valuable addition to our Marburg Collection 2.0 and to the iGEM Registry of Parts. It services users of our chassis and other cyanobacterial strains with a useful tool for genomic modifications but it also contributes a shell that can be used to modify any other model organism as well.