Difference between revisions of "Team:Lethbridge/Results"
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| − | <h1> Purification of Arc | + | <h1> Purification of Full-Length Arc and Minimal Arc Gag</h1> |
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<p class="f14">Lorem Ipsum is simply dummy text of the printing and typesetting industry. Lorem Ipsum has been the industry's standard dummy text ever since the 1500s, when an unknown printer took a galley of type and scrambled it to make a type specimen book. It has survived not only five centuries, but also the leap into electronic typesetting, remaining essentially unchanged. It was popularised in the 1960s with the release of Letraset sheets containing Lorem Ipsum passages, and more recently with desktop publishing software like Aldus PageMaker including versions of Lorem Ipsum.</p> | <p class="f14">Lorem Ipsum is simply dummy text of the printing and typesetting industry. Lorem Ipsum has been the industry's standard dummy text ever since the 1500s, when an unknown printer took a galley of type and scrambled it to make a type specimen book. It has survived not only five centuries, but also the leap into electronic typesetting, remaining essentially unchanged. It was popularised in the 1960s with the release of Letraset sheets containing Lorem Ipsum passages, and more recently with desktop publishing software like Aldus PageMaker including versions of Lorem Ipsum.</p> | ||
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<h2>Full-Length Arc</h2> | <h2>Full-Length Arc</h2> | ||
| − | <center><img src="https://static.igem.org/mediawiki/2018/ | + | <center><img src="https://static.igem.org/mediawiki/2018/7/7a/T--Lethbridge--ArcFullTEM.png" alt="TEM Full" style="width: 600px"></center> |
<p>TEM image (60,000X magnification) of negatively-stained self-assembled full-length Arc PNCs. Scale bar = 200um.</p> | <p>TEM image (60,000X magnification) of negatively-stained self-assembled full-length Arc PNCs. Scale bar = 200um.</p> | ||
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<h2>P22 coat proteins</h2> | <h2>P22 coat proteins</h2> | ||
| − | <center><img src="https://static.igem.org/mediawiki/2018/ | + | <center><img src="https://static.igem.org/mediawiki/2018/4/48/T--Lethbridge--TEM-P22-NoSP.png" alt="TEM P22" style="height: 500px"></center> |
| − | <p>TEM image (40,000X magnification) of negatively-stained self-assembled P22 capsids in the absence of scaffolding proteins. Scale bar = | + | <p>TEM image (40,000X magnification) of negatively-stained self-assembled P22 capsids in the absence of scaffolding proteins. Scale bar = 500um.</p> |
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<h2>P22-SP-Cas9</h2> | <h2>P22-SP-Cas9</h2> | ||
| − | <center><img src="https://static.igem.org/mediawiki/2018/ | + | <center><img src="https://static.igem.org/mediawiki/2018/0/0e/T--Lethbridge--TEM-P22-SP-Cas9.jpg" alt="TEM P22-SP-Cas9" style="height:500px"></center> |
| − | <p>TEM image (40,000X magnification) of negatively-stained self-assembled P22 capsids with the SP-Cas9 cargo. Scale bar = | + | <p>TEM image (40,000X magnification) of negatively-stained self-assembled P22 capsids with the SP-Cas9 cargo. Scale bar = 200um.</p> |
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</div> | </div> | ||
Revision as of 16:46, 16 October 2018
Purification of Full-Length Arc and Minimal Arc Gag
Lorem Ipsum is simply dummy text of the printing and typesetting industry. Lorem Ipsum has been the industry's standard dummy text ever since the 1500s, when an unknown printer took a galley of type and scrambled it to make a type specimen book. It has survived not only five centuries, but also the leap into electronic typesetting, remaining essentially unchanged. It was popularised in the 1960s with the release of Letraset sheets containing Lorem Ipsum passages, and more recently with desktop publishing software like Aldus PageMaker including versions of Lorem Ipsum.
Transmission Electron Microscopy
Transmission electron microscopy (TEM) confirmed the presence of spherical nanoparticles in both the full-length M. musculus Arc protein and minimal Arc Gag protein samples. Even in the absence of RNA cargo, the Arc-derived PNCs were capable of self-assembly in vitro. In both samples, the average PNC diameter was approximately 30nm +/- 10nm.
Full-Length Arc
TEM image (60,000X magnification) of negatively-stained self-assembled full-length Arc PNCs. Scale bar = 200um.
Minimal Arc Gag
TEM image (100,000X magnification) of negatively-stained self-assembled minimal Arc Gag PNCs. Scale bar = 100um.
TEM also confirmed the presence of spherical nanoparticles in the P22 protein sample (without added scaffolding proteins; SPs) and P22 + SP-Cas9 protein samples. In the absence of the scaffolding protein, P22 capsid proteins were capable of self-assembly but with a significant variation in size and stability (many collapsed structures were evident in the sample). The P22 + SP-Cas9 PNCs had a much more consistent architecture but had a diameter of 120 +/- 5nm, which is approximately twice that of typical P22 virus-like particles. The 1:1 capsid to cargo ratio may have caused overloading of the PNCs, which caused an increase in overall size.
P22 coat proteins
TEM image (40,000X magnification) of negatively-stained self-assembled P22 capsids in the absence of scaffolding proteins. Scale bar = 500um.
P22-SP-Cas9
TEM image (40,000X magnification) of negatively-stained self-assembled P22 capsids with the SP-Cas9 cargo. Scale bar = 200um.
Mass Spectrometry
Lorem Ipsum is simply dummy text of the printing and typesetting industry. Lorem Ipsum has been the industry's standard dummy text ever since the 1500s, when an unknown printer took a galley of type and scrambled it to make a type specimen book. It has survived not only five centuries, but also the leap into electronic typesetting, remaining essentially unchanged. It was popularised in the 1960s with the release of Letraset sheets containing Lorem Ipsum passages, and more recently with desktop publishing software like Aldus PageMaker including versions of Lorem Ipsum.
Minimal Arc Gag
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P22 Capsid
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Analytical Ultracentrifugation of Minimal Arc Gag
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Cell Culture Transfection with Minimal Arc Gag
Lorem Ipsum is simply dummy text of the printing and typesetting industry. Lorem Ipsum has been the industry's standard dummy text ever since the 1500s, when an unknown printer took a galley of type and scrambled it to make a type specimen book. It has survived not only five centuries, but also the leap into electronic typesetting, remaining essentially unchanged. It was popularised in the 1960s with the release of Letraset sheets containing Lorem Ipsum passages, and more recently with desktop publishing software like Aldus PageMaker including versions of Lorem Ipsum.
Purification of P22 and SP-Cas9
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Collaboration with the University of Calgary iGEM Team
Lorem Ipsum is simply dummy text of the printing and typesetting industry. Lorem Ipsum has been the industry's standard dummy text ever since the 1500s, when an unknown printer took a galley of type and scrambled it to make a type specimen book. It has survived not only five centuries, but also the leap into electronic typesetting, remaining essentially unchanged. It was popularised in the 1960s with the release of Letraset sheets containing Lorem Ipsum passages, and more recently with desktop publishing software like Aldus PageMaker including versions of Lorem Ipsum.
Improved Parts
We characterized 15 parts from the 2017 Lethbridge iGEM team's "Next Vivo" project. Please visit the Improved Parts page for links to the new registry pages and all related data.