Team Marburg has built a collection containing the promoter, RBS, terminator, tags, and
different ORIs. Promoters in this collection have a high homogeneity, however, which may
cause homologous recombination in the V.n due to its rapid growth rate. What’s more,
the copy number spectrum of ORIs is narrow and regulation methods are lack due to the poor
measurement of inducible promoters.
This year, taking up-elements, insulator, special 5’-UTR, inducible promoters, artificially
designed promoters, and a tunable ORI into consideration, we expended the Marburg
collection, hoping to develop a collection for strict and
subtle regulation (SSR collection).
1. Up-elements: In SSR collection, the up-elements is rich in PAM
5’-TBN-3’ which is compatible with CasΦ-2 (CasΦ for short below) , so the expression of
genes in our circuit can be regulated by CRISPRa.
2. Insulators: The insulator is a type of self-cleavage ribozyme that
could cut itself as quickly as transcription reaches the 5’ of insulator. In this iGEM
project, we use RiboJ widely.
3. Twelve generators based on Marionette collection: All of them are from
E.coli marionette collection, and most of them show fantasy cooperative in
E.coli. Here, we introduced and modified them in Vibirio.natriegens.
4. Artificial designed promoters: All of them are from E.coli
marionette collection, and most of them show fantasy cooperative in E.coli. Here, we
introduced and modified them in Vibirio.natriegens.
5. Tunable ORI:After characterizing basic ORIs in Vibirio.natriegens,
we found ColE1 performed well, so we engineering it into Tet-ColE1 so as to control the copy
number in a wide range