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<img align="center" src="https://static.igem.org/mediawiki/2016/1/12/RepairTrans.png" alt = "repair" /> | <img align="center" src="https://static.igem.org/mediawiki/2016/1/12/RepairTrans.png" alt = "repair" /> | ||
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− | <h4 align="center"> Although the researchers were able to add an inhibitor to their system to resist this enzyme there were still some issues with this approach. There was only a 50% chance of their edited strand being chosen as the template in cellular mismatch repair and the desired edit occurring. | + | <h4 align="center"> Although the researchers were able to add an inhibitor to their system to resist this enzyme there were still some issues with this approach. There was only a 50% chance of their edited strand being chosen as the template in cellular mismatch repair and the desired edit occurring. Also, the researchers could only perform a C to U edit, so the capabilities were limited. </h4> |
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<h4 align="center"> Based off of this, we started to think about editing the mRNA. But is targeting the RNA with CRISPR/Cas9 possible? </h4> | <h4 align="center"> Based off of this, we started to think about editing the mRNA. But is targeting the RNA with CRISPR/Cas9 possible? </h4> | ||
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− | <h4 align="center"> A paper published in 2014 showed that it is possible to target RNA with CRISPR through the use of a PAMmer. This is a short oligonucleotide that is separate from Cas9 but helps to guide the system there and activates nuclease | + | <h1 align="center"> Targeting RNA </h1> |
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+ | <h4 align="center"> A paper published in 2014 showed that it is possible to target RNA with CRISPR through the use of a PAMmer. This is a short oligonucleotide that is separate from Cas9 but helps to guide the system there and activates nuclease to performs the cleavage. Through the use of a PAMmer, dCas9 was also shown to be able to target mRNA. </h4> | ||
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<img align="center" src="https://static.igem.org/mediawiki/2016/6/63/PammerTrans.gif" alt = "pammer" /> | <img align="center" src="https://static.igem.org/mediawiki/2016/6/63/PammerTrans.gif" alt = "pammer" /> | ||
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<h3 align="center"> <b> Advantages </b> </h3> | <h3 align="center"> <b> Advantages </b> </h3> | ||
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− | <h4 align="center"> There are many advantages to targeting mRNA. If any mistakes happen, they will not be permanent because the DNA is not being edited. The system will be regulated with doxycycline, and we can double the | + | <h4 align="center"> There are many advantages to targeting mRNA. If any mistakes happen, they will not be permanent because the DNA is not being edited. The system will be regulated with doxycycline, and we can double the possible number of edits. Not only will we be able to use APOBEC1 and perform C to U edits, but we will also be able to use the editing enzyme ADAR, which performs A to I edits exclusively in RNA. Inosine (I) is a nucleotide that is common in brain pathways and is read by the ribosome as a G. By combining all of these concepts together, we developed a system to target single nucleotide changes in RNA. You can read more about the specifics of our system in our <a href="https://2016.igem.org/Team:WPI_Worcester/Design"> Design </a>.</h4> |
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+ | <h3 align="center"><b>References</b></h3> | ||
+ | <h4>O'Connell, M.R., et al.(2014).Programmable RNA recognition and cleavage by CRISPR/Cas9.<i>Nature</i>.516:263-266.<br><br> | ||
+ | Komor, A.C., et al.(2016).Programmable editing of a target base in genomic DNA without double-stranded DNA cleavage.<i> Nature</i>533:420-424.<br><br> | ||
+ | Nelles DA et al. (2016).Programmable RNA Tracking in Live Cells with CRISPR/Cas9.<i>Cell</i>.165:488-496.<h4> | ||
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Latest revision as of 23:53, 19 October 2016