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Fluorescent Biosensors for Measuring Autophagic Flux

Posted by Beth Kenkel on Jan 22, 2019 9:41:01 AM

Autophagy (Greek for “self-eating”) is a process by which cytoplasmic material, including organelles, are targeted to lysosomes for degradation. Autophagy is a dynamic process which involves autophagosome synthesis, delivery of materials to be degraded to the lysosome, and degradation of autophagic substrates inside the lysosome. Historically, methods for studying autophagy focused on counting the number of autophagosomes. This approach, however, has inherent limitations because it turns a dynamic process into a static measurement and it provides limited information about what materials or organelles are being targeted for autophagy. The development of several fluorescent autophagy reporters now allows for the measurement of autophagic flux, or the changes in autophagic activity, and are a more reliable indicator of autophagic activity. The aim of this post is to provide an overview of four autophagy biosensors currently available from Addgene.

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Topics: Plasmid Technology, Fluorescent Proteins

15 Years of Addgene: The Top 15 Plasmids

Posted by Jennifer Tsang on Jan 8, 2019 8:54:52 AM

15 years of plasmid sharing has certainly taken us on many adventures...From moving office locations three times, to opening our UK office in 2014, to starting our viral vector service in 2016...we’re excited to help scientists share their reagents with the scientific community.

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Topics: Plasmid Technology, Scientific Sharing, Inside Addgene

Plasmid addiction systems: from bacterial toxins to molecular biology tools

Posted by Jennifer Tsang on Nov 1, 2018 8:35:58 AM

Members of the bacterial world produce an assortment toxins to claim territory or kill competing microorganisms, but did you know bacteria also produce substances toxic to themselves?

These toxic substances are part of toxin-antitoxin systems that are widely present in bacteria. They consist of a toxin which can affect a variety of cellular processes and an antitoxin that suppresses the toxin’s activity. The key to these systems is that the toxin is stable while the antitoxin is unstable, meaning that the cells must continually produce antitoxin to avoid cell death.

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Topics: Plasmid Technology, Plasmid Elements, Plasmid Cloning

Stabilized Bacterial Promoters: Constant Gene Expression at any Copy Number

Posted by Jennifer Tsang on Sep 4, 2018 8:53:28 AM

Researchers express genes of interest from plasmids in order to study gene function or to engineer cells for specific purposes. Unfortunately, plasmid copy numbers vary within cell populations and over time resulting in variable gene expression that can impact observed phenotypes. Factors such as the growth medium, growth temperature, and growth rate can all impact plasmid copy number in a cell.

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Topics: Plasmid Technology, Plasmid Elements, Synthetic Biology

Finding nucleic acids with SHERLOCK and DETECTR

Posted by Alyssa Cecchetelli on Aug 30, 2018 8:28:06 AM

Sensitive and specific nucleic acid detection is crucial for clinical diagnostics, genotyping, and biotechnological advancements. Current 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. Recently scientists have utilized CRISPR-Cas9 protein variants, Cas13, and Cas12a, to develop simple, portable, and inexpensive platforms to reliably detect nucleic acids at the atomolar level.

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Topics: Plasmid Technology, Genome Engineering, CRISPR

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