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History of CRISPR Cas - A tale of survival and evolution

Posted by Margo R. Monroe on Nov 25, 2013 5:12:00 PM

All organisms share an innate goal to survive. This past year, scientists hijacked survival tactics of prokaryotes to deliver the technological biological blockbuster known as the CRISPR (clustered regularly interspaced palindromic repeats) Cas (CRISPR associated genes) system. This popular genome engineering tool offers flexibility, multiplexibility, and ease of use. In order for this technology to survive the diverse demands of the biotech field, let's look at how prokaryotes originally utilized CRISPR/Cas as a powerful and adaptive defense strategy against life-threatening viruses.

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Topics: Plasmid Technology, Genome Engineering, CRISPR

Let There Be LITE Plasmids

Posted by Kendall Morgan on Nov 21, 2013 10:56:00 AM

When neuroscience graduate student Silvana Konermann first entered Feng Zhang’s lab at MIT, the use of customizable DNA-binding domains based on transcription-activator-like effectors (TALEs) as anchors for genome engineering applications was still very new. Now, Konermann and her colleagues including Zhang and Mark Brigham have taken the technology to another level with the addition of two light-sensitive ingredients - CRY2 and CIB1 – which they borrowed from Arabidopsis thaliana. The results are light-inducible transcriptional effectors (LITEs) designed to bind specific genes and turn them on or off, literally at the flip of a (blue) light switch.

Zhang’s team devised the plasmids now available in Addgene’s repository for use in neurons, both in culture and in living brain tissue. “The brain is such an amazing and dynamic organ,” Konermann explained. “It helps us adapt to influences in our environments. We are able to react and learn. All of this requires genes to be regulated dynamically.”

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Topics: Plasmid Technology, Genome Engineering, Optogenetics

Tag Your Favorite Yeast Genes with Ease

Posted by Julian Taylor-Parker on Nov 19, 2013 9:37:00 AM

Homologous recombination is the process by which nearly all domains of life repair genomic damage, specifically double strand breaks. Researchers have long taken advantage of this natural process to integrate protein tags into the genomes of S. cerevisiae and S. pombe. The protocol is surprisingly simple, requiring only a PCR product containing the modifying sequence flanked by approximately 50 base pairs of sequence homologous to the chromosomal site of insertion. The linear PCR product is introduced into the cell by direct transformation. A given insert will typically contain both a protein modification sequence and a selectable gene product for isolation of successful transformants.

Addgene distributes several ready-to-use, modular plasmids, combining fluorescent tags, epitope tags, protease sites, and selection markers. These are especially useful in protein complex studies where tagging of multiple protein products is desired, as multiple selection markers can ensure that all desired tags have been integrated. Simply design your amplification primers with the desired targeting homology—in frame, of course—and start tagging!

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Topics: Plasmid Technology, Hot Plasmids, Genome Engineering, Yeast

Addgene’s New Way to Help You Share Science

Posted by Caroline LaManna on Nov 14, 2013 11:10:00 AM

When I got the job of Outreach Scientist at Addgene, my first thought was “I get to talk to scientists from around the world making breakthrough discoveries!” I knew right then I had found my dream job, because one thing I learned in grad school was that scientists have amazing stories to share.

I spend a lot of time telling scientists about Addgene’s repository, and explaining how to deposit and request plasmids, but I also spend a lot of time listening.

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Topics: Scientific Sharing, Inside Addgene

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