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Adeno-associated Viruses (AAVs) for Genome Editing

Posted by Tyler Ford on Mar 27, 2018 9:32:43 AM

Guest blogger Todd Waldman, Professor at Georgetown University, contributed to this post.

Adeno-associated viruses (AAVs) make fantastic gene delivery vehicles for episomal gene expression and are particularly useful for gene delivery to the nervous system. For many years they have also been used to enhance the efficiency of genome editing. In this post we'll walk through a variety of ways you can use AAVs to improve your genome editing experiments (with and without targeted nucleases).

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Topics: Hot Plasmids, Viral Vectors

New Neuroscience Tool: The SF-iGluSnFr Glutamate Sensor

Posted by Tyler Ford on Mar 8, 2018 9:00:00 AM

In a previous blog post we discussed how fluorescent proteins can be used to construct biosensors, biological tools that monitor processes or detect molecules. Here we’ll be diving into the details surrounding SF-iGluSnFr, a recently upgraded biosensor designed to detect glutamate.

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Topics: Hot Plasmids, Fluorescent Proteins

Plasmids 101: Secondary Nanobody Toolbox

Posted by Beth Kenkel on Feb 27, 2018 9:04:41 AM

Western blots. ELISAs. Immunofluorescence. What do all of these techniques have in common? They all typically require secondary antibodies, frequently of the mouse or rabbit variety. While antibodies certainly aren’t “broken,” their production does require continued animal sacrifice. Could there be an alternative method for immunodetection? Enter the Görlich lab and their anti-mouse and -rabbit IgG secondary nanobodies toolbox. Nanobodies are like tiny antibodies which work just as well, if not better, than antibodies for all of the above listed molecular techniques, but they can also be expressed in bacteria and extracted with common protein purification methods. Read on to learn more about nanobodies and how their structure and function compare to IgG antibodies, as well as how to produce them for use in your lab.

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Topics: Hot Plasmids, Plasmids 101

In Vivo Biotinylation of Bacterial Fusion Proteins

Posted by Guest Blogger on Jan 25, 2018 9:09:35 AM

This post was contributed by guest blogger Jon Backstrom, a biochemist in the Vanderbilt Eye Institute and Tonia Rex's lab.

A common strategy to determine the binding kinetics of a purified protein involves immobilization on a solid support. This allows washing away of unbound material to calculate the amount of bound ligand (after subtracting out non-specific binding). Historically, glutathione-S-transferase (GST) fusion proteins have been immobilized on a reduced glutathione matrix. The advantage of a fusion protein is the efficient purification of an already immobilized target protein. The disadvantage is that the GST moiety, which forms dimers, may influence binding kinetics of the target ligand. Another important consideration is whether the affinity of an experimental protein-ligand interaction approaches that of GST-glutathione.

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Topics: Plasmid Technology, Hot Plasmids, Techniques

Top Requested AAV of 2017: pmSyn1-EBFP-CRE

Posted by Tyler Ford on Jan 17, 2018 9:57:12 AM

We began distributing ready-to-use virus preps through our viral service in late 2016 and requests are still pouring in! While our lentiviral service is going strong, the AAV service has shown incredible growth this year. pAAV-hSyn-DIO-hM4D(Gi)-mCherry was the top requested AAV prep for the 2nd year running, and you can learn more about this useful, DREADD-containing AAV here. But the top requested AAV that became available in 2017 is pmSyn1-EBFP-Cre from Hongkui Zeng’s lab. This AAV has had over 150 orders since coming online!

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Topics: Plasmid Technology, Hot Plasmids, Viral Vectors

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