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... colonies to isolate the monomer Sometimes a plasmid can exist in a bacterial population as a mixture ...growing the transformed bacterial culture into the late stationary phase. Growing the culture at a lower...detection methods described above can help ensure the isolation of the monomeric version of the plasmid. Linearize...ligate the plasmid If you are having difficulty isolating a plasmid monomer from a plasmid sample, or if... CP, Carnes AE. Generic plasmid DNA production platform incorporating low metabolic burden seed-stock ...

...long been extremely useful in the lab – they are relatively small and easy to work with, making them the ...much genetic and physiological data have been accumulated over the years using them. Indeed, the future...work is bright as it is now easier than ever to manipulate the mouse genome using CRISPR/Cas9. Similar ...would be great if we could just go in and quickly manipulate individual mouse genes and study their function...successful in my first attempt at using CRISPR? Later on, I found out that my experience was shared by...s41467-017-02706-7 Wang H, Yang H, Shivalila CS, Dawlaty MM, Cheng AW, Zhang F, Jaenisch R (2013) One-Step...

...thought that it was necessary to develop a single platform to connect talented professionals (who migrated...and their successes. Although all mentorship relationships are valuable and highly recommended, a mentor...mentor creates a closer and more approachable relationship for the mentee, where (s)he feels truly understood..., mentors and mentees often developed close relationships with a level of sympathy that is difficult to...confidence. These improvements, combined with the relationships fostered through the IMP have motivated many...-Harvard Medical School, where she studies age-related macular degeneration. She is also a blogger who...

...caused this glowing phenomenon wasn’t until much later when, in the 1960s, Osamu Shimomura studied the ...Washington state. Using these samples, he was able to isolate two proteins from the jellyfish’s photoorgans; ... absorbs that blue light and glows green. In the late 1980s, another researcher, Douglas Prasher, got ...expressed GFP in exogenous organisms (E. coli, and later C. elegans) for the first time. After Chalfie’s ...fluorescent proteins like GFP really are. In this compilation we’ll take a closer look at how they’re used,...

...editing system is regulated not only temporally with doxycycline, but it can also be regulated spatially with...these base editors Spatially and temporally regulated AAV-based genome editing In 2016, Jonathan Ploski... mediated genome editing system that could be regulated by doxycycline. Now, they’ve developed another... that CRISPR efficiency can be controlled by manipulating complementarity between the gRNA and target....

...react and learn. All of this requires genes to be regulated dynamically.” To understand how that works in ...other brain functions, Konermann needed tools to manipulate the expression of specific genes in the brain...and genetically target them to specific cell populations. They demonstrated use of the LITEs to control...transcription in as little as 30 minutes after light stimulation. Under those optogenetic controls, gene expression...chromophores flavin-adenine dinucleotide and methyltetrahydrofolate, which are universally abundant. Second...

...interests are very clear, I had no idea how to translate them into real world careers. I figured this internship...but this never happened. I actually started at a later date and worked entirely virtually. Stepping into...world, I was worried about how my ideas would be translated through Zoom calls and Google docs. I pondered...Canva, and I was able to create cute and on-brand templates for these posts. Some of recent Meet an ... departments of the company. I asked questions relating to their roles at Addgene. I had a great time ...

...plastic you’ve saved. Repurposing regulated medical waste Regulated medical waste (RMV), which we collect...environmental impact of those processes? Who even regulates them? You may be surprised to learn that it hasn...Environmental Protection Agency, which started making regulations such as preventing manufacturers from dumping...and reused materials such as glass pipettes or inoculation loops. While single-use is now a popular theme...

...application we were looking at, magnetic isolation of circulating tumor cells, we found that the binding...Oxford is dedicated to generating new tools to manipulate biology based on molecular features found in ...the SpyTag/SpyCatcher system, based on a protein isolated from Streptococcus pyogenes that locks itself ...the context of cancer? Howarth: The capture of circulating tumor cells (CTCs) is going to have a big impact...

...on antibiotic plates.  Importantly, because the bacteria from which plasmids are isolated grow quickly ...easily make large amounts of plasmid to manipulate and use in later work.     How do scientists use plasmids...plasmids? Generally, scientists use plasmids to manipulate gene expression in target cells. Characteristics...

...antibiotic resistance marker. After transformation, plate the cells on medium containing the corresponding...contains a kanamycin resistance cassette, you would plate the cells on kanamycin-containing agar. Only cells... are auxotrophic for specific amino acids. When plated on media lacking the essential amino acid, the ... a toxic metabolite in gram-negative bacteria. Plating on sucrose medium will select for cells that contain...gene is used for negative selection on 5% sucrose plates. Exchange of the sacB gene for the gene of interest...

...specific, programmable platform well-suited for various innovative biological and translative technologies. Addgene...only deactivate the Type I-F system and are not translatable to the other CRISPR subtypes. Still, the large...Mali et al. in January 2013 (7, 8). Nine months later, more than 1500 publications describe work to improve...enabling access to multiple plasmids used for various platform applications. For more information on CRISPR/Cas...

...with regard to compliance with health authority regulations,” Baradaran says.  Baradaran’s path to science...is an Addgene depositor. KSQ uses a CRISPR-Cas9 platform to systematically test the function of all 20,... a genetic screen to identify the factors that regulate cell division. And that really got me hooked on...lentiviral microRNA-based system for polymerase II-regulated RNA interference. In the first two months since...also interested to see Addgene expand the sharing platform to other reagents in biology. Luk H. Vandenberghe...

...establish a next-generation sequencing (NGS) QC platform that allows us to provide scientists with complete...launching our viral service in 2016, we adapted this platform to accommodate adeno-associated viral vector (...ligation and tagmentation require a double-stranded template.  Using viral genome sequencing to verify the ...double-stranded species that could be used as a template for downstream applications (Berns, 1972). Using...the Addgene blog Read all of our viral vectors related blog posts Read our blog post on AAV titers Browse...

...pAAV.hSyn-FLEX.iGABASnFR.F102G) is noticeably dimmer than iGABASnFR, and accumulates in the endoplasmic reticulum, but exhibits larger...GlyR IRES EGFP Chimeric channels for neuronal manipulation 119742  AAV5  AAV SYN PSAM4 GlyR IRES EGFP...EGFP Chimeric channels for neuronal manipulation Recombinases Plasmid Serotype Name 50363  AAV5 .... It is useful for activity imaging of large populations of densely labeled neurons because background...  It is useful for activity imaging of large populations of densely-labeled neurons because background...

... of the time because of relationships. It’s much harder to build relationships when you are more desperate...your hand at a video presentation? Most video platforms make it easy to record yourself. You can watch...experiences to share. The conversation builds a relationship for the future and the 1:1 format is good for...you will make good progress on both building relationships and thinking about future career options. 5....

...activity of two populations of neurons at once. Addgene spoke to Boyden about this latest optogenetics ...compete against each other. Getting two-color stimulation has been a holy grail of the field. The problem...selective for different ions. Right now, we can modulate electricity. If you could selectively let in calcium...Independent Optical Excitation of Distinct Neural Populations. Nathan Klapoetke et al. Nature Methods. 2014...

...site-specific recombinases, regulatory elements, developmental regulators, and lots of viral components... as Pichia pastoris). If your goal is protein isolation, then the MoClo Pichia Kit will get you started...developed to study the basic biology of and gene regulation in the opportunistic pathogen Acinetobacter baumannii... Molecular Tools to Study Gene Expression and Regulation in Acinetobacter Baumannii.” ACS Synthetic Biology.... “A Unified Multi-Kingdom Golden Gate Cloning Platform.” Scientific Reports, vol. 9, no. 1, Springer ...

...switched on and off reversibly. This ability to modulate fluorescence allows scientists to observe single... (T203H) (1). wtGFP normally contains a mixed population of neutral (protonated) and anionic (deprotonated...T203H mutation in PA-GFP generates a chromophore population that is more neutral, causing PA-GFP to display...with blue light (1). Similar to PA-GFP, scientists later developed red fluorescent PA-FPs by performing several...photons achievable with photoswitchable dyes. Correlative light and electron microscopy (CLEM) is an imaging...Ryoko, Hideaki Mizuno, and Atsushi Miyawaki. "Regulated fast nucleocytoplasmic shuttling observed by reversible...

...vector production, assembled AAV vector particles accumulate inside the producer cells and purification protocols...gradients. The current purification protocols for isolating exo-AAVs rely on differential centrifugation steps...Figure 2). Since the development of the exo-AAV isolation protocol by Maguire and colleagues, other researchers...et al., 2017), and exo-AAV8 expressing human coagulation factor IX in liver in a mouse model of Haemophilia...et al. 2018). The structure of exosome AAVs is related to the use of microvesicles as a vehicle for drug...