This year Team Fudan-TSI presents you with a toolbox for our in vivo sequence-specific continuous mutagenesis system. This toolbox includes promoters, RBSs, terminators, CDSs and other regulatory parts which undergo careful design and optimization. The Moloney Murine Leukemia Virus Reverse Transcriptase (MMLV-RT), which is new to the registry. Cre recombinase are the most important components of our system for which we have put emphasis on the construction and characterization. Also, our improved Lac repressor protein is fully equipped for regulating protein expression.
Specifically, the parts that we are presenting this year include but are not limited to these following features:
1) We attached Cre with different degradation tags, which could be used according to user’s interest to achieve optimal recombination efficiency.
2) We placed MMLV-RT under different IPTG-inducible promoters, which provides a range of different steady-state expression levels for various experimental purposes.
3) We included additional regulatory sequences (e.g. native primer binding site and poly-purine tract) required for the initiation and completion of reverse transcription.
4) To eliminate self-excision of Cre and promote recombination efficiency, we included a set of incompatible loxP sites.
5) We provide testing plasmids for system verification and optimization.
| Bronze Requirement | BBa_K1021005 BBa_E0040 |
| Silver Requirement | BBa_K3257071 BBa_K3257072 BBa_K3257073 |
| Gold Requirement | BBa_C0012 → BBa_K3257045 |
| Our Best Basic Part | BBa_K3257042 |
| Our Best Composite Part | BBa_K3257101 |
| Our Part Collection |
In summary, our part collection provides a complete set for assembly, test, and optimization of continuous mutagenesis in different prokaryotic hosts.