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The PAM Requirement and Expanding CRISPR Beyond SpCas9

Posted by Joel McDade on Aug 20, 2020 10:30:00 AM

Originally published Nov 12, 2015 and last updated Aug 20, 2020.

Cas9 can be used to modify any desired genomic target provided that (1) the sequence is unique compared to the rest of the genome and (2) the sequence is located just upstream of a Protospacer Adjacent Motif (PAM sequence). The 3-5 nucleotide PAM sequence serves as a binding signal for Cas9 and this sequence is a strict requirement for Cas9-mediated DNA cleavage.

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Topics: CRISPR, Cas Proteins

CRISPR 101: RNA Editing with Cas13

Posted by Mary Gearing on Jul 31, 2020 8:30:00 AM

Originally published Nov 30, 2017 and updated Jul 31, 2020.

Cas13 enzymes are quickly becoming major players in the CRISPR field. Just a year after Feng Zhang’s lab identified Cas13a (C2c2) (Abudayyeh et al., 2016) as a RNA-targeting CRISPR enzyme, they adapted Cas13b for precise RNA editing (Cox et al., 2017). This new system, termed REPAIR (RNA editing for programmable A to I (G) replacement) is the first CRISPR tool for RNA editing. Two years after that, the lab published a paper on an RNA editor that allows C to U edits (RESCUE). We’ll walk through how these tools were developed and potential ways you can use it in your research.

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Topics: CRISPR, CRISPR 101, Cas Proteins

SARS-CoV-2/COVID-19 Detection Methods Based on CRISPR/Cas

Posted by Guest Blogger on May 5, 2020 9:15:00 AM

This post was contributed by Shravanti Suresh from Iowa State University.

Since its appearance, SARS-CoV-2 has spread to almost every part of the world manifesting as a full-fledged pandemic. Containing the spread of this virus has become an utmost priority for countries around the world and to do so, the WHO recommends one strategy: testing, tracking, and social distancing.

With countries like South Korea, one of the earlier epicenters of the outbreak ultimately flattening the curve, it has become evident that widespread testing is crucial in controlling this pandemic. Currently, the CDC uses RT-qPCR tests to diagnose COVID-19 and some serological tests to determine past exposure. However, the limited availability of reagents and equipment and the long turnaround times, have led researchers to turn to other technologies like CRISPR.

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Topics: CRISPR, Cas Proteins, COVID-19

Finding nucleic acids with SHERLOCK and DETECTR

Posted by Alyssa Cecchetelli on Apr 16, 2020 9:00:09 AM

Originally published Aug 30, 2018 and updated April 16, 2020.

Sensitive and specific nucleic acid detection is crucial for clinical diagnostics, genotyping, and biotechnological advancements. Many methods of nucleic acid detection however, either lack the sensitivity or the specificity to detect nucleic acids at low concentrations and/or are too expensive, time-consuming, and complex to use outside of standard laboratories. In the case of the COVID-19 pandemic, qPCR can be used to diagnose the presence of SARS-CoV-2 RNA, but inadequate access to reagents and equipment has become a bottleneck.

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Topics: CRISPR, Cas Proteins, Other CRISPR Tools, COVID-19

Behind-the-scenes of the Isolation of the Thermostable IgnaviCas9 From a Yellowstone Hot Spring

Posted by Christina Mork on Nov 12, 2019 9:00:00 AM

In 2008 the Quake Lab at Stanford University became interested in exploring biological dark matter – large tracts of the microbial tree of life that remained unexplored. Using new single-cell sequencing approaches, the lab was able to eliminate the need for axenic (pure) laboratory cultures to study these microbes. From 16S rRNA sequencing, hot springs were known to be diversity hotspots containing abundant biological dark matter and so the lab organized a sampling trip to Yellowstone National Park (YNP).

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Topics: CRISPR, Cas Proteins

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