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Team:Warwick/Parts
From 2014.igem.org
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<h2>MS2 Box</h2> | <h2>MS2 Box</h2> | ||
| - | <p> | + | <p>The MS2 box acts as a binding |
| + | |||
| + | site for the MS2 coat protein, formed | ||
| + | |||
| + | downstream in the replicon, which | ||
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| + | represses translation hence preventing | ||
| + | |||
| + | exponential growth of the replicon.</p> | ||
</div> | </div> | ||
<div id="pop4" class="popbox"> | <div id="pop4" class="popbox"> | ||
<h2>Neomycin Resistance</h2> | <h2>Neomycin Resistance</h2> | ||
| - | <p> | + | <p>This was included in order to select |
| + | |||
| + | for the cells which had been | ||
| + | |||
| + | successfully transfected with the | ||
| + | |||
| + | replicon. This is commonly used in | ||
| + | |||
| + | human cell studies and allows survival | ||
| + | |||
| + | of eukaryotic cells in the prescence of | ||
| + | |||
| + | geneticin.</p> | ||
</div> | </div> | ||
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Revision as of 21:45, 16 October 2014
5' Promoter
This is derived from the 5i UTR of the HCV virus strain 1b isolate Con1. The secondary structure of this sequence acts as a binding site and initiates replication of the system by RdRp. It also includes the first 16 amino acids of the first gene of HCV as this has been shown to increase the efficacy of binding of the RdRp to the 5' promoter.
IRES
This acts as an initiation for eukaryotic ribosomes and begins translation of the following protein sequence. We compared two different IRESs: the classical EMCV IRES used in many papers,which has been shown to be compatible with replicons and the NKRF IRES derived from the 3’UTR of the mammalian NF-kappaB repressing factor, which we also chose to test because during investigation regarding the efficacy and strength of the EMCV IRES, the NKRF derived IRES was shown to be 30-fold more efficient however had never been previously used in Huh7.
MS2 Box
The MS2 box acts as a binding site for the MS2 coat protein, formed downstream in the replicon, which represses translation hence preventing exponential growth of the replicon.
Neomycin Resistance
This was included in order to select for the cells which had been successfully transfected with the replicon. This is commonly used in human cell studies and allows survival of eukaryotic cells in the prescence of geneticin.
Aptazyme
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siRNA
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MS2 Coat Protein
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P2A
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RdRp
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3' Promoter
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