Difference between revisions of "Team:Calgary/Results"
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<h3 class="infosubtitle">Results Overview</h3> | <h3 class="infosubtitle">Results Overview</h3> | ||
<br> | <br> | ||
| − | <p style="text-indent: 0px"> | + | <p style="text-indent: 0px"> uCalgary was able to compare the effectiveness of 7 different sgRNA candidates at directing CRISPR/Cas9 cleavage. These results are pseudo-quantitative, providing the relative rate of indel formation which can be used as a surrogate measure for cleavage activity and NHEJ.</p> |
| − | + | ||
| − | + | <p> chart </p> | |
| − | + | ||
| − | + | ||
<br> | <br> | ||
| − | <p style="text-indent: 0px"> | + | <p style="text-indent: 0px"> uCalgary was not able to sequence confirm the targeted insertion of an FRT site using CRISPR/Cas9. However, attempted integration showed significant indel formation, demonstrating that Cas9 was successfully cleaving the targeted DNA. The issue lay with the incorporation of template DNA through homology directed repair. </p> |
| − | + | <br> | |
| − | + | <p> image </p> | |
| − | + | <br> | |
| − | + | <p> Despite no sequencing data confirming insertion, the amount of DNA sequenced and analyzed did not cover the entire cell population. This leaves the possibility that integration was successful, albeit at extremely low efficiency. To further support this hypothesis, uCalgary found that a small number of transfected cells successfully grew in puromycin-containing cell media. Puromycin resistance was placed promoter-less on a plasmid which was designed to enter the genome through Flp recombinase. Thus, growth in puromycin indicates the possibility that not only did some cells successfully receive an FRT insert from CRISPR, but they also had our desired plasmid integrated into their genome using our recombinase system at the FRT site. We are optimistic about this qualitative measure of success and hope to expand upon these results in the brief time before the Jamboree. | |
| − | + | <br> | |
| + | <p> image </p> | ||
<br> | <br> | ||
<p style="text-indent: 0px">Lorem ipsum dolor sit, amet consectetur adipisicing elit. Rerum sit perferendis eum delectus odit vero saepe, | <p style="text-indent: 0px">Lorem ipsum dolor sit, amet consectetur adipisicing elit. Rerum sit perferendis eum delectus odit vero saepe, | ||
Revision as of 07:07, 17 October 2018
RESULTS
Results Overview
uCalgary was able to compare the effectiveness of 7 different sgRNA candidates at directing CRISPR/Cas9 cleavage. These results are pseudo-quantitative, providing the relative rate of indel formation which can be used as a surrogate measure for cleavage activity and NHEJ.
chart
uCalgary was not able to sequence confirm the targeted insertion of an FRT site using CRISPR/Cas9. However, attempted integration showed significant indel formation, demonstrating that Cas9 was successfully cleaving the targeted DNA. The issue lay with the incorporation of template DNA through homology directed repair.
image
Despite no sequencing data confirming insertion, the amount of DNA sequenced and analyzed did not cover the entire cell population. This leaves the possibility that integration was successful, albeit at extremely low efficiency. To further support this hypothesis, uCalgary found that a small number of transfected cells successfully grew in puromycin-containing cell media. Puromycin resistance was placed promoter-less on a plasmid which was designed to enter the genome through Flp recombinase. Thus, growth in puromycin indicates the possibility that not only did some cells successfully receive an FRT insert from CRISPR, but they also had our desired plasmid integrated into their genome using our recombinase system at the FRT site. We are optimistic about this qualitative measure of success and hope to expand upon these results in the brief time before the Jamboree.
image
Lorem ipsum dolor sit, amet consectetur adipisicing elit. Rerum sit perferendis eum delectus odit vero saepe, dignissimos aspernatur et libero quisquam minima soluta a suscipit tempora dolores non aliquid ratione? Lorem ipsum dolor, sit amet consectetur adipisicing elit. Sunt excepturi quod, doloribus et asperiores similique tempora, mollitia possimus doloremque officia deleniti eius aut dolorum fuga reiciendis adipisci esse quisquam quae.
Why CRISPR?
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Lorem ipsum dolor sit amet consectetur adipisicing elit. Temporibus rerum vel eius ut dolore, ab obcaecati officiis modi porro, sunt deleniti, consequatur assumenda asperiores aliquid recusandae tenetur neque quae suscipit! Lorem ipsum dolor sit amet consectetur adipisicing elit. Optio a quam iusto quo, nesciunt odit fuga, similique aspernatur veritatis nemo commodi libero nobis magnam necessitatibus, quidem maiores error debitis minima. Lorem ipsum dolor sit amet consectetur adipisicing elit. Quam ipsum consequatur, deserunt assumenda odio natus. Quis, ea dolor! Voluptas dolore, facere cum illo sunt consectetur nam a soluta optio perferendis.
Lorem ipsum dolor sit, amet consectetur adipisicing elit. Rerum sit perferendis eum delectus odit vero saepe, dignissimos aspernatur et libero quisquam minima soluta a suscipit tempora dolores non aliquid ratione? Lorem ipsum dolor, sit amet consectetur adipisicing elit. Sunt excepturi quod, doloribus et asperiores similique tempora, mollitia possimus doloremque officia deleniti eius aut dolorum fuga reiciendis adipisci esse quisquam quae.