Difference between revisions of "Team:Jilin China/Design"
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| + | <p>This year, our project aims to design a thermosensor toolkit, which could sense different temperature and hold different intensity, sensitivity. After three versions of the upgrade, our toolkit consist of four types of thermosensors, which areheat-inducible, heat-repressible, cold-inducible and cold-repressible RNA-based thermosensors. We name this as SynRT toolkit, Synthetic RNA-based thermosensors, which principle is similar to natural RNA thermosensors. Naturally occurring RNA thermomsensors exhibit complex secondary structures which are believed to undergo a series of gradual structural changes in response to temperature shifts. However, they lack standardization and only respond to temperatures in a narrow range，which impends application process[1]. Therefore, we focus on more rational and engineering design after understanding the principles of natural RNA thermosensors response. | ||
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| + | <p>RNA-based thermosensors mediate the temperature-controlled access of the ribosome to the SD sequence, because of the SD sequence is buried in a stem-loop structure, which is melting as temperatures rising[2].Stem-loop intramolecular base pairing is a pattern that can occur in single-stranded DNA or, more commonly, in RNA. A base pair (bp) is a unit consisting of two nucleobases bound to each other by hydrogen bonds, such as A-U or G-C.Based on natural RNA thermosensors’ stem-loop sequence, we would like to modify the stem-loop region in order to amplify our tool kits with different melting temperature. With the increasing of base pairing or hydrogen bonds in the stem-loop structure, the free energy required for conformation transitions would be higher. According to this, we could know qualitatively that increasing base pairing or GC content could increase the melting temperature, which means the temperature for unfolded mRNA moleculars occupying the half of mRNA moleculars. But we need further quantitative parameters of thermosensors, such like the detailed free energy of conformation transition, before our wet experiment, so we apply mfold web server (Mfold) to help us engineer our designs. Mfold describes a number of closely related software applications available on the World Wide Web for the prediction of the secondary structure of single stranded nucleic acids[3]. RNA secondary structure is predicted by energy minimization using nearest neighbor energy parameters. After prediction and selection, desired RNA-based thermosensors have been obtained and then they have been measured in E.coil. | ||
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Revision as of 12:18, 15 October 2018
DESIGN
Toolkits Design
- Overview
- Heat-inducible Thermosensors
- Heat-repressible Thermosensors
- Cold-inducible Thermosensors
- Cold-repressible Thermosensors
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title
This year, our project aims to design a thermosensor toolkit, which could sense different temperature and hold different intensity, sensitivity. After three versions of the upgrade, our toolkit consist of four types of thermosensors, which areheat-inducible, heat-repressible, cold-inducible and cold-repressible RNA-based thermosensors. We name this as SynRT toolkit, Synthetic RNA-based thermosensors, which principle is similar to natural RNA thermosensors. Naturally occurring RNA thermomsensors exhibit complex secondary structures which are believed to undergo a series of gradual structural changes in response to temperature shifts. However, they lack standardization and only respond to temperatures in a narrow range，which impends application process[1]. Therefore, we focus on more rational and engineering design after understanding the principles of natural RNA thermosensors response.
RNA-based thermosensors mediate the temperature-controlled access of the ribosome to the SD sequence, because of the SD sequence is buried in a stem-loop structure, which is melting as temperatures rising[2].Stem-loop intramolecular base pairing is a pattern that can occur in single-stranded DNA or, more commonly, in RNA. A base pair (bp) is a unit consisting of two nucleobases bound to each other by hydrogen bonds, such as A-U or G-C.Based on natural RNA thermosensors’ stem-loop sequence, we would like to modify the stem-loop region in order to amplify our tool kits with different melting temperature. With the increasing of base pairing or hydrogen bonds in the stem-loop structure, the free energy required for conformation transitions would be higher. According to this, we could know qualitatively that increasing base pairing or GC content could increase the melting temperature, which means the temperature for unfolded mRNA moleculars occupying the half of mRNA moleculars. But we need further quantitative parameters of thermosensors, such like the detailed free energy of conformation transition, before our wet experiment, so we apply mfold web server (Mfold) to help us engineer our designs. Mfold describes a number of closely related software applications available on the World Wide Web for the prediction of the secondary structure of single stranded nucleic acids[3]. RNA secondary structure is predicted by energy minimization using nearest neighbor energy parameters. After prediction and selection, desired RNA-based thermosensors have been obtained and then they have been measured in E.coil.
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