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The PAM Requirement and Expanding CRISPR Beyond SpCas9

Posted by Joel McDade on Nov 12, 2015 10:30:00 AM

This post was updated on Dec 5, 2017.

Diverse genomes and genomic targets require a variety of tools to engineer them effectively. Since the discovery and engineering of dCas9, the CRISPR toolbox has expanded to include a variety of natural and engineered Cas proteins. Read on to learn how these tools can be used to expand CRISPR's reach to new genomic loci. 

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Topics: Genome Engineering, CRISPR

Inntags: Innovative Protein Epitope Tagging

Posted by Mary Gearing on Nov 10, 2015 10:30:00 AM

First described in the 1980s, protein tags are now one of the most useful items in a scientist’s toolbox. As we’ve covered in Plasmids 101, tags can help you determine localization of a protein of interest, purify it, or determine its expression level without the need for a custom antibody. There is one major caveat - a tag may interfere with protein localization and/or function, so each tagged protein must be tested carefully to ensure it retains the attributes of the native protein. Since this process takes a lot of time and energy, Martí Aldea and collaborators have created a set of “innocuous tags” (inntags) less likely to alter a protein’s properties.

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Topics: Hot Plasmids, Plasmid Elements

Building Global Connections with the International Mentorship Program USA-EUROPE

Posted by Guest Blogger on Nov 6, 2015 10:30:00 AM

This post was contributed by guest bloggers and IMP organizers Rosario F. Godino, Rocío López-Diego, & Zafira Castano Corsino.

When someone says “internationalization” young hearts often shake with the fear of uncertainty. However, internationalization, the ability to cross country boundaries for both professional and personal development, is essential to excelling in the modern world. Talent knows no boundaries and should not be confined by birth country or financial means. Instead, talent should be realized through equal opportunities for all regardless of origin. It is therefore essential that internationalization become a reality available to all youth, our future, around the world.

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Topics: Career, Networking, Career Readiness

The CRISPR Software Matchmaker: A New Tool for Choosing the Best CRISPR Software for Your Needs

Posted by Guest Blogger on Nov 3, 2015 10:30:00 AM

This post was contributed by guest blogger Cameron MacPherson at the Institut Pasteur

CRISPR Software and the Piñata Effect

Two years ago I was a part of a group (Biology of Host-parasite Interactions, Institut Pasteur, Paris) that changed genome editing in the malaria community for the better (Nat. Biotechnol., 2014). Given the timing, it shouldn’t be a surprise that the CRISPR system was involved. Today, that same laboratory enjoys a successful edit rate of over 90% in their work editing the genome of Plasmodium falciparum (the parasite that causes malaria). I attribute their success to technical expertise, thoughtful single guide RNA (sgRNA) design, and the abnormally low GC content of the Plasmodium falciparum genome. To put this last point into perspective, the Plasmodium falciparum genome contains only 0.66 million targetable NGG PAM sites whereas the human genome has about 300 million. With such a sparsely targetable genome, off-targeting is less of a worry and on-targeting likely more efficient. 

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Topics: Genome Engineering, CRISPR

Plasmids 101: A Brief History of Plasmids and an Improved eBook!

Posted by Marcy Patrick on Oct 29, 2015 10:30:00 AM

Bioblasts? Plasmagenes? In the 1940s and 50s, scientists were working to understand genetic cytoplasmic factors that could be transferred between cells. At the time, these extranuclear agents of heredity were thought of as everything from parasites, to symbionts, to genes and the labels applied to them were vague or contradictory, owing in part to the fact that very little was known about the role these factors played within an organism. 

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Topics: Plasmids 101, Plasmid Cloning

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