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<div align="center"><p >Fig.8 The comparison between experimental data and simulation data</p></div> | <div align="center"><p >Fig.8 The comparison between experimental data and simulation data</p></div> | ||
− | <a href="https://2018.igem.org/Team:OUC-China/miniToe" style="font-size: 25px; color: blue"> To see more details </a> <br /><br /> | + | <a href="https://2018.igem.org/Team:OUC-China/miniToe" style="font-size: 25px; color: blue; text-decoration:none;"> To see more details </a> <br /><br /> |
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Revision as of 19:45, 16 October 2018
Overview
The aim of our project is to develop a better post-transcriptional regulation strategy and use it in monocistron and polycistron. We build models to design and predict our work.
miniToe —— a better transcriptional regulate strategy
To achieve a better post-transcriptional regulation strategy, we design a system which is composed of an RNA endoribonuclease (Csy4) and an RNA module named miniToe. We model to describe the dynamics of the miniToe system and point out the way to achieve different regulation level. The ODE and molecular dynamics are two main tools to explore it. We use the ODE to describe the reaction curve and the molecular dynamics give some explanations to experimental data.
Below you can follow the several questions we point out to have a better understanding of model work and the miniToe system. We will discuss some structures of Csy4 in different stage (Q1), some structures of miniToe system in different stage (Q2), the reaction order and some keys of miniToe system (Q3), the simulation of ODE model (Q4), some significant symbol in molecular dynamics (Q5) and the way to different regulation level (Q6).
Q1 : What does the structure of Csy4?
Fig.1 The structure of Csy4 without hairpin bound (PDB ID: 4AL5, resolution 2.0 A)
Fig.2 The structure of Csy4 with hairpin bound (PDB ID: 4AL5, resolution 2.0 A)
Q2 : What does the structure of miniToe structure?
1. A cis-repressive RNA (crRNA) to serve as translation suppressor by pairing with RBS as the critical part of miniToe structure.
2. A Csy4 site as a linker between cis-repressive RNA and RBS, which can be specifically cleaved upon Csy4 function.
3. A CRISPR endoribonuclease Csy4.
Fig.2-1 is the secondary structure of miniToe.
Fig.2 The structure of miniToe.
Fig.4 The precursor complex of wild-type Csy4
Fig.5 The product complex of wild-type Csy4
Q3 : What is the reaction order of miniToe system?
Fig.6 The working process of miniToe system
(1)The miniToe structure is produced and accumulated.
(2)The Csy4 is produced with IPTG induced.
(3)The Csy4 binds to the miniToe structure and form the Csy4-miniToe complex
(4)The Csy4 cleave the special site and divide the miniToe structure into two parts: the Csy4-crRNA complex and the mRNA of sfGFP.
(5)The sfGFP is produced.
From the description above, we can get four key problems in our system to make sure that our system can work successfully:
(1)Does the Csy4 dock correctly with the miniToe structure (hairpin)?
(2)How about the ability of binding between the Csy4 and miniToe structure (hairpin)?
(3)How about the ability of cleavage between the Csy4 and miniToe structure (hairpin)?
(4)Does cis-repressive RNA release from the RBS?
Q4 : How about the simulation result of the ODE model?
According to the work process we build an ODEs model and simulate our miniToe system for 30h, the result can be seen in the Fig.7.
Fig.7 The dynamics of sfGFP by model prediction
Fig.8 The comparison between experimental data and simulation data
Q5 : What does the structure of Csy4?
Fig.1 The structure of Csy4 without hairpin bound (PDB ID: 4AL5, resolution 2.0 A)
Fig.1 The structure of Csy4 without hairpin bound (PDB ID: 4AL5, resolution 2.0 A)
Q6 : What does the structure of Csy4?
Fig.1 The structure of Csy4 without hairpin bound (PDB ID: 4AL5, resolution 2.0 A)
Fig.1 The structure of Csy4 without hairpin bound (PDB ID: 4AL5, resolution 2.0 A)