Difference between revisions of "Team:SCUT China/Overview"

Parts Overview

We have registered 31 basic parts (BBa_ K3100001-BBa_K310003) and 41 composite parts (BBa_K3100100-BBa_K3100140), all of which is BioBrick RFC10 or Type IIS compatible.

BBa_K3100001 to BBa_K3100010 is different strength T7 promoter which is from the research of Amar Ghodasara et al.^[1]. Toehold Switch is prokaryotic riboregulators^[2], that activate gene expression in response to cognate RNAs( Trigger RNAs) with arbitrary sequences. We have added Toehold Switch A&C and their Trigger DNAs (Toehold Switch B&D is from previous part number range). BBa_K3100100-BBa_K3100139 are composite parts which are the combination of 4 different Triggers and 10 unique T7 promoters. Through specific design of Fusion Sites, these 40 parts will be randomly combined by Golden Gate assembly. Up to 4 different Triggers driven by 10 unique promoters are expressed from one plasmid and finally a pool called VerProS with a storage capacity of 10000 is obtained. VerProS pool can simultaneously optimize up to four genes in a system. Particularly, this versatile library can be applied to fast optimization in different systems without having to build ad hoc libraries, which can greatly reduce manpower and costs.

We have added 4 acid tolerant factors (BBa_K3100017- BBa_K3100020) and have tested their influence on the acid tolerance of E.coli. What’s more, BBa_K3100140 is a composite part combined of 4 acid tolerant factor and 4 Toehold Switch. In our Project, we use the VerProS pool for fine regulation of these 4 acid tolerant factors.

As T7 is the most well-known inducible promoter with high transcriptional strength ^[3], we want to construct and improve a T7 promoter library which contains more T7 promoters with different strength. Therefore, the library can help our VerProS system to achieve more fine-grained regulation. Site-saturation mutagenesis was conducted to obtain the T7 mutants. BBa_K3100022-BBa_K3100031 is the different strength T7 promoters which we have constructed and improved.

[1] Ghodasara A, Voigt CA. (2017) Balancing gene expression without library construction via a reusable sRNA pool.
[2] Green AA, Silver PA, Collins JJ, Yin P. (2014) Toehold switches: de-novo-designed regulators of gene expression.
[3] Nie, Z., Luo, H., Li, J. et al. Appl Biochem Biotechnol (2019). https://doi.org/10.1007/s12010-019-03113-y