Difference between revisions of "Part:BBa K2904050"

Revision as of 12:53, 13 October 2019

aTc inducible sfGFP regulated by modular Adda riboswitch containing Tuner A

Background of 2019 OUC-China' project——RiboLego

Due to context-dependent performance and limited dynamic range, the widespread application of riboswitches is currently restricted. By replacing its original ORF with a new one, the structure of an aptamer domain can be subtly disrupted, resulting in a loss of ligand response. So riboswitch is still not be considered as a ‘plug and play' device. To tackle these problems, our project focuses on a standardized design principle to be used for modular and tunable riboswitch. The modular riboswitch we defined consists of the original riboswitch, Stabilizer and Tuner. Stabilizer can protect the structure of riboswitch from damage while Tuner can reduce the expression probability of fusion protein and make improvement of riboswitch function. We test our design principle in different riboswitches including three kinetic switches: Adda riboswitch, Btub riboswitch, cobalamin biosensor, and one thermodynamic switch: FourU riboswitch. What's more, three different kinds of GOI is used including sfGFP, YFP, and mRFP1. The good results show the high universality of our design principles. We hope our guideline can easily be appied by future teams. we design a toolkit named miniToe family focused on translational regulation, which is composed of a RNA endoribonuclease, Csy4 and a RNA module (hairpin). In our project, the cleavage function of Csy4 releases a cis-repressive RNA module (crRNA, paired with RBS) from the masked ribosome binding site (RBS), which subsequently allows the downstream translation initiation. A Ribosome Binding Site (RBS) is an RNA sequence to which ribosomes can bind and initiate translation.
The sfGFP was regulated by the Adda riboswitch containing the original Adda riboswitch, Stabilizer and Tuner A. Sequence and Features