View source for Part:BBa K3806020


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<partinfo>BBa_K3806020 short</partinfo>

The TU Delft iGEM 2021 project <html><a href="https://2021.igem.org/Team:TUDelft"><b>AptaVita</b></a></html> looks at developing a modular, quantitative, and accessible rapid diagnostic test for vitamin deficiencies. In this project, vitamin-specific aptazyme biosensors are engineered through a novel <html><i>in vitro</i></html> evolution method, <html><i>De novo</i></html> Rapid <html><i>In vitro</i></html> Evolution of RNA biosensors (DRIVER) [1]. Range <HTML><a href="https://parts.igem.org/Part:BBa_K3806017" target="_blank"><b>BBa_K3806017</b></a></HTML> - <HTML><a href="https://parts.igem.org/Part:BBa_K3806021" target="_blank"><b>BBa_K3806021</b></a></HTML> is set of primers designed to assist the DRIVER selection process.

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<span class='h3bb'>Sequence and Features</span>
<partinfo>BBa_K3806020 SequenceAndFeatures</partinfo>



==Usage and Biology==
<HTML><h3>DRIVER</h3></html>
Starting with a high-diversity library of potential biosensors, DRIVER makes use of a regeneration strategy within a directed-evolution procedure. This grants the selection of sequences that present a higher level of cleavage in the absence of ligand and a lower level of cleavage in the presence of ligand. During rounds in which the ligand(s) are present (also referred to as uncleavage selection rounds), the uncleaved RNA molecules preserve an unmodified 5’ prefix sequence from the previous round. These sequences are PCR amplified using this prefix and a specific suffix as primers. During rounds in which the target ligand(s) are absent (cleavage rounds), RNA molecules that undergo self-cleavage are regenerated by addition of a different 5’ prefix. Afterwards, these sequences are PCR amplified using the newly added prefix and the specific suffix as primers. By iteratively applying this process, each round selectively enriches the desired subset of sequences from the library. Eventually, RNA sequences whose cleavage is regulated by the target ligand(s) dominate the population. 


<HTML><u><b>BBa_K3806020 in DRIVER</b></u></html>

To allow for prefix replacement in cleavage selection rounds, two prefix sequences are required, named X prefix and Z prefix. <HTML><a href="https://parts.igem.org/Part:BBa_K3806020" target="_blank"><b>BBa_K3806020</b></a></HTML>  is used as the forward primer to amplify sequences with the W prefix.

Sequence: 5’ AATTTAATACGACTCACTATAGGGAAACAAACAAAGCTG 3’


==References==
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[1]Townshend, B., Xiang, J. S., Manzanarez, G., Hayden, E. J. and Smolke, C. (2021). A multiplexed, automated evolution pipeline enables scalable discovery and characterization of biosensors. Nat Commun, 12, 1437.
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===Functional Parameters===
<partinfo>BBa_K3806020 parameters</partinfo>
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