<div dir="ltr">Justin, <div>I set up a zoom call for Tuesday at 1 pm and sent out an invite. Can you please share this invite with your student and ask him to join as well?</div><div>Thanks! -Borries</div></div><br><div class="gmail_quote"><div dir="ltr" class="gmail_attr">On Fri, Jul 7, 2023 at 7:56 AM Pahara, Justin (AAFC/AAC) <<a href="mailto:justin.pahara@agr.gc.ca">justin.pahara@agr.gc.ca</a>> wrote:<br></div><blockquote class="gmail_quote" style="margin:0px 0px 0px 0.8ex;border-left:1px solid rgb(204,204,204);padding-left:1ex"><div class="msg-7819905569742603557">
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<p class="MsoNormal">Good Morning Borries, <u></u><u></u></p>
<p class="MsoNormal"><u></u> <u></u></p>
<p class="MsoNormal">Good question – we had not planned any purification at this point. The reason being we are ultimately interested in aptamers that bind to different types of macromolecules within the lysate (protein, carbs, etc). If we run a purification
it will bias toward a particular system. I was hoping we could do a one pot process during this prototype experiment.
<u></u><u></u></p>
<p class="MsoNormal"><u></u> <u></u></p>
<p class="MsoNormal">It might be possible to label the aptamer with both biotin and a fluorophore so that we can pull out the aptamers with beads… what do you think? I was hoping to start with a simple experiment and add complexity (and expense) as needed,
so what do you think is the simplest starting point??<u></u><u></u></p>
<p class="MsoNormal"><u></u> <u></u></p>
<p class="MsoNormal">A call next week sounds great – how about Tuesday? 1pm?<u></u><u></u></p>
<p class="MsoNormal"><u></u> <u></u></p>
<p class="MsoNormal"><u></u> <u></u></p>
<p class="MsoNormal">Best wishes,<u></u><u></u></p>
<p class="MsoNormal">Justin.<u></u><u></u></p>
<p class="MsoNormal"><u></u> <u></u></p>
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<p class="MsoNormal"><b>From:</b> Borries Demeler <<a href="mailto:demeler@gmail.com" target="_blank">demeler@gmail.com</a>> <br>
<b>Sent:</b> Tuesday, July 4, 2023 3:02 PM<br>
<b>To:</b> Pahara, Justin (AAFC/AAC) <<a href="mailto:justin.pahara@AGR.GC.CA" target="_blank">justin.pahara@AGR.GC.CA</a>>; <a href="mailto:demelerlab@biophysics.uleth.ca" target="_blank">demelerlab@biophysics.uleth.ca</a><br>
<b>Subject:</b> Re: update on AUC experiment<u></u><u></u></p>
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<p class="MsoNormal">Hi Justin, <u></u><u></u></p>
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<p class="MsoNormal">this sounds like a great start! So many opportunities for applying multi-wavelength AUC. I was wondering what level of purification, if any, has been done on these complexes. Potentially a sucrose gradient could be used to concentrate some
of the complexes. It would be wonderful if we could get the labeled molecules by themselves as controls, both to determine their sedimentation coefficients and their absorbance spectra when they are pure so we can distinguish them from the rest of the proteins
in your lysate, are there any purifications planned?<u></u><u></u></p>
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<p class="MsoNormal"><u></u> <u></u></p>
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<p class="MsoNormal">Perhaps we could set up a Zoom call next week with the rest of the lab members and your ULeth student?<u></u><u></u></p>
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<p class="MsoNormal"><u></u> <u></u></p>
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<p class="MsoNormal">-Borries<u></u><u></u></p>
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<p class="MsoNormal"><u></u> <u></u></p>
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<p class="MsoNormal">On Tue, Jul 4, 2023 at 1:11 PM Pahara, Justin (AAFC/AAC) <<a href="mailto:justin.pahara@agr.gc.ca" target="_blank">justin.pahara@agr.gc.ca</a>> wrote:<u></u><u></u></p>
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<p class="MsoNormal">Good Afternoon Borries,
<u></u><u></u></p>
<p class="MsoNormal"> <u></u><u></u></p>
<p class="MsoNormal">I hope you’re enjoying a good start to summer
<span style="font-family:"Segoe UI Emoji",sans-serif">😊</span><u></u><u></u></p>
<p class="MsoNormal"> <u></u><u></u></p>
<p class="MsoNormal">I want to provide a quick update about the research we discussed earlier. We have begun to prototype our first SELEX experiment where we’ve designed a ssDNA aptamer library and
we plan to run a mock experiment with whole cell lysate from E. coli expressing RFP or CFP and then try absorbance/fluorescence ultracentrifugation with you to identify fraction(s) of Aptamer-RFP. We could also use chromoproteins if that is better and you’d
prefer AUC. If possible it would be great to not have the aptamer fluorescently tagged for this first experiment.<u></u><u></u></p>
<p class="MsoNormal"> <u></u><u></u></p>
<p class="MsoNormal">Our initial aptamer library is 90 nts long with 50 Ns, so ~30 kDa and the RFP/CFP is about the same, so we’re looking at around ~60 kDA for single DNA/protein interaction.<u></u><u></u></p>
<p class="MsoNormal"> <u></u><u></u></p>
<p class="MsoNormal">What do you think? Might we be able to try it out later this month? The summer student who is working on the project is a ULeth student and so he could do most of the work if your
team is busy! <u></u><u></u></p>
<p class="MsoNormal"> <u></u><u></u></p>
<p class="MsoNormal">Best,<u></u><u></u></p>
<p class="MsoNormal">Justin.<u></u><u></u></p>
<p class="MsoNormal"> <u></u><u></u></p>
<p class="MsoNormal"> <u></u><u></u></p>
<p class="MsoNormal"><b><span lang="EN-CA" style="font-size:10pt">Dr. Justin Pahara</span></b><u></u><u></u></p>
<p class="MsoNormal"><b><span lang="EN-CA" style="font-size:10pt"> </span></b><u></u><u></u></p>
<p class="MsoNormal"><b><span lang="EN-CA" style="font-size:10pt">Research Scientist and Project Lead</span></b><u></u><u></u></p>
<p class="MsoNormal"><span lang="EN-CA" style="font-size:10pt"> </span><u></u><u></u></p>
<p class="MsoNormal"><span lang="EN-CA" style="font-size:10pt">Nanotechnology (Biotic Stresses and Adaptation)</span><u></u><u></u></p>
<p class="MsoNormal"><span lang="EN-CA" style="font-size:10pt">Agriculture and Agri-Food Canada / Government of Canada<br>
</span><a href="mailto:justin.pahara@agr.gc.ca" target="_blank"><span lang="EN-CA" style="font-size:10pt;color:rgb(5,99,193)">justin.pahara@agr.gc.ca</span></a><u></u><u></u></p>
<p class="MsoNormal"><span lang="EN-CA" style="font-size:10pt"> </span><u></u><u></u></p>
<p class="MsoNormal"><span lang="FR-CA" style="font-size:10pt">Nanotechnologie (Adaptation et Contraintes Biotiques)</span><span lang="FR-CA"><u></u><u></u></span></p>
<p class="MsoNormal"><span lang="FR-CA" style="font-size:10pt">Agriculture et Agroalimentaire Canada / Gouvernement du Canada</span><span lang="FR-CA"><u></u><u></u></span></p>
<p class="MsoNormal" style="margin-bottom:12pt"><a href="mailto:justin.pahara@agr.gc.ca" target="_blank"><span lang="FR-CA" style="font-size:10pt;color:rgb(5,99,193)">justin.pahara@agr.gc.ca</span></a><u></u><u></u></p>
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