We narrowed to 897 results for: ANT

...by guest bloggers Erik L. Snapp and Lindsey M. Costantini. "You underestimate the power of the Dark Side...non-fluorescent, and misfolded FP-fusions on quantitative imaging experiments and cell physiology in general... the cytoplasm? Or more troubling, what if a significant fraction of the fusion protein molecules fail...population is not readily apparent and can confound quantitative imaging experiments or even negatively impact...fusion based on your experimental needs. Equally important, before cloning you must consider whether and ...resident proteins within these compartments undergo significant post-translational modifications including glycosylation...misfolded dark FP fusions including erroneous quantitation of fusion protein levels. It remains unclear...

...The FLAG antibody has been well characterized and the system allows you to add this same antibody to any...known to have good antibodies. The results look almost identical. Addgene: What advantages does your system...established method called ChIP-seq. ChIP-seq relies on antibodies to recognize and bind transcription factors directly...the transcription factor. It also gives cells antibiotic resistance to neomycin so that you can select...of the proteins inside a cell have a commercial antibody against them. There’s also a question about how... solved in this CRISPR system with a universal antibody. It’s an epitope not found in human or other cells...transcription factors, but there are many reasons you might want a system like this. It’s actually surprising to ...

...Salary –not relevant to most postdocs. Commercial successes in the performing arts- not relevant at all. ...options discussed in this post benefit from the applicant thinking about the process early, keeping good... minimum (8 U.S. Code § 1184 - Admission of nonimmigrants). The foreign worker must possess at least a... cap subject H-1B (last year less than 30% of applicants received sponsorship) then he or she can start... facility management, collaboration management, grant administration, clinical operations, and so on. ...1B petitioners are exempt from the cap if the applicant works at institutions of higher education, a related...US Immigration officers use to determine if an applicant qualifies for the National Interest Waiver. The...

... our ready-to-use recombinant antibodies service, part of the Neuroscience AntiBody Open Resource (NABOR...library of antibody educational resources, including our new Antibodies 101 eBook, the Antibody Guide, and...could use the same principles to provide recombinant antibodies, another plasmid-based product that, with...front of us. So in our last celebratory moment, we wanted to walk through some of our favorite milestones...designing a website users could easily navigate. This meant we could grow accessibility alongside our repository...so easy!” — Nina Jain, Harvard Medical School. We want to give a special thanks to the early-adopter scientists...beginners” on r/labrats, Reddit.  Part of that work meant facilitating the Material Transfer Agreement process...

...are numerous GFP variants. Here I use the name ‘GFP’, which refers to mEGFP. This variant was made by introducing...contributed by guest blogger Joachim Goedart, an assistant professor at the Section of Molecular Cytology...jellyfish GFP (AvGFP) to create enhanced GFP (EGFP), a variant with improved brightness (Tsien, 1998). Another...necessary to generate the strictly monomeric EGFP variant, mEGFP (Zacharias et al., 2002). GFP needs oxygen...that generates the chromophore (Tsien, 1998). Importantly, this reaction requires molecular oxygen and ...Cranfill et al., 2016 and FPbase.org). A number of variants remain fluorescent under acidic conditions, with...N-terminus, but this hardly reduces its size. The substantial size of GFP may interfere with the activity or...

... community which is really important to her. Addgenie Shreya Vedantam with our resident dinosaur...school. QC Scientist Alyssa Cecchetelli knew she wanted to get her PhD. “Going back to school was a pretty...in Addgene since he started, said, “the thing I wanted more of [after undergrad] was to learn and speak...may open doors for you, but depending on where you want to go in your career you may not need those doors...her first job at Addgene as an Order Processing Assistant. After a few months, she left school when she ...Technician. “I was so tired of being in school so I just wanted to work,” she said. “School will always be there...mean you’ve failed. “I got a job as a research assistant in a lab at the university that I went to. Making...

...Notably, the SpRY variant didn’t display higher off-target effects than SpCas9, a pleasant surprise since... PAM requirement, and how these proteins are advantageous in today’s genome editing landscape. The need...anywhere within the RNA will do. Similarly, if you want to knock out a gene, a frameshift mutation introduced...that don’t utilize NGG   There are plenty of Cas9 variants which utilize alternative PAMs. These alternative...looking for more pan-genome PAM flexibility, you’ll want to consider one of several Cas9s that recognize ...compatibility. This alternative PAM requirement adds a significant amount of flexibility compared to a strict single-compatibility...rounds of engineering, they developed the SpRY Cas9 variant (Walton, et. al, 2020). SpRY uses NRN PAMs and ...

...recombination software looks for and quantifies the presence of recombinants in recombinase-dependent sequences...experiments to confirm their findings. But no scientist wants to repeat an experiment because of poor reagent ...production process? These questions are especially important when producing multiple vectors in parallel because...serotype of the packaged DNA and look for potential contaminants. The analysis is a 2-step process using Geneious...total reads are most likely to correspond to the contaminant. Each contig is then BLASTed against the NCBI...contaminated. For AAV-68544, the best match for the contaminant is Addgene plasmid #44362. Method validation,... were easily able to identify AAV-44632, the contaminant we mixed into the original AAV sample at 5%. ...

...neurons. Their soma-targeted variant SomaFRCaMPi also performed substantially better than other soma-localized...manipulation transplantation (AMT), CRISPR-modified adipocytes (fat cells) are transplanted as a mass that...months we highlight some of the new plasmids, antibodies, viral preps, and more in the repository through...function, but these connections are exquisitely important in the brain. To better understand how astrocytes... proposed, researchers have shown that these transplanted mitochondria can exert their powers for better...use quite different designs, leading to unique advantages. While most red GECIs have been derived from ... and signal response upon Ca2+ binding. Most importantly, it doesn’t display the problematic photoswitching...

... to reassess what I wanted to do and so I applied for a role in Industry. I wanted to get "a real job"...then, and still feel now, that it is important to take advantage of the fact that science is a global ...staff hired in anticipation of the contract suddenly have less work to do than was anticipated and there can... Sutton, located on the edge of London to gain relevant experience. I applied for this summer position...abroad in rural Africa Once this position ended, I wanted to explore another area which I was really passionate...Procedures (SOP’s) and every item I tested was constantly tracked with barcodes and spreadsheets. It was...to be in the lab doing experiments  without the constant fear of failure because I knew the tests worked...

... tissue temperature proximal to the implanted fibers. As important as its capabilities are the practical...Sands, J., Zhu, W., Tabet, A., Manthey, M., Allen, H., Loke, G., Antonini, M.-J., Rosenfeld, D., Park, ...gut-brain axis (admittedly for self-serving reasons – I want to understand my obsession with the combination ...connection to study on a molecular level. The brain is constantly influenced by signals it receives from the entire...included in them for downstream control – this is important later! Next, additional components are added -... be, as well as other features necessary for implantation in brain tissue. The gut, on the other hand...hand, was the wild west for fiber implants. Due to the soft nature of the intestines and the requirement...

...to aid in the expression and purification of recombinant proteins. Tags for stability and solubility ...hurdles to overcome in order to overexpress a recombinant protein? It is not generally in a cell’s best...glutathione S-transferase (GST), which can be fused to recombinant proteins for one-step purification with glutathione...uncharacterized protein, or a protein for which a good antibody has not been developed (and just because your ...your protein has a commercially available antibody, that doesn’t mean it’s a good one), then your first step... an affinity tag to it. The FLAG, hemaglutinin antigen (HA), and c-myc tags have been the workhorses of...depend on your application (see table below). The antibodies available for these tags really are good and ...

...repression using Cas9 Don’t want to permanently modify the genome but still want to affect gene expression...post was originally written by Joel McDade and significantly updated in 2022 by Susanna Stroik.  The advent...your experimental target, SpCas9 and its enhanced variants, eSpCas9 and HypaCas9, are great choices. Check...regions of the genome and consider using a Cas9 variant with higher specificity (eSpCas9 and HypaCas9) ...added bonus of low off-target effects. The SpG variant, discovered by the Kleinstiver group has a PAM ...Further, this group also characterized a second Cas9 variant, SpRY, which cuts at NRN PAMs with high efficiency...that the desired edit will be performed on all relevant bases within the editing window, for example all...

...editing by APOBEC-Cas9 Though the ACE reporter quantifies APOBEC-Cas9 efficiency in real-time, it relies...Harris Lab created a panel of eGFP reporters to quantify the on-target DNA editing efficiency of APOBEC-Cas9...would restore eGFP fluorescence, making it easy to quantify single base editing efficiency by dividing the... et al., 2016). For TREE they engineered a BFP variant that undergoes conversion to GFP after being targeted...deaminase base DNA editor. Specifically the BFP mutant (BFPH66) contains a histidine at amino acid 66....changes the histidine to a tyrosine generating a GFP variant (GFPY66) that has a shifted emission spectra. Thus...the test by infecting HEK-293 cells with the BFP variant, a BE4am-Cas9 base editor, and sgRNA vectors for...

...for every application. Disadvantages include their duplicated (i.e., redundant) genome, their long three-month..., including investigations into environmental pollutants and health, embryo growth, brain function, and...straightforward husbandry and ability to produce abundant, externally fertilized, transparent progeny (Figure...visualize expression data in useful ways. Zebrafish mutants In the 1990’s, two revolutionary large-scale random... mutagenesis screens produced ~1,500 zebrafish mutants that are still used in labs across the globe (Nüsslein-Volhard...which I will come back to) is the one-eyed pinhead mutant, which regulates a developmental pathway implicated... in congenital heart defects and cancer. These mutants are used extensively to study development and disease...

...Westwood) Dodders are parasitic plants that steal all their food from other plants and are unusual enough to...efficiency to reduce the number of plants you need to grow to get a recombinant because this process is expensive...between gene function and phenotypes in diverse plants, animals, microbes, viruses, and fungi for which...next project. Gymnotiformes and Mormyridae (Jason Gallant) Weakly electric fish are excellent systems to ...are now studied in 40 labs as a great model for plant adaptation and speciation. Their phenotypes are ...great for studying parenting strategies and make important ecological indicators since species live in water...Aiptasia - algae Breviolum minutum symbionts are important models for solutions to the coral reef crisis....

...to infect your cells of interest.   Advantages Disadvantages Can carry large transgenes (>8 kb) Potential...is an important feature for your experiment; it must be chosen carefully. Often researchers want a simple...particles into the culture supernatant and you can generally harvest the supernatant containing viruses 24h...cells. These properties make lentiviral vectors fantastic options for delivering shRNA, CRISPR/Cas9 components... cells, it was obvious that its safety was a disadvantage particularly for clinical development. In addition...using them.   Regarding biosafety, it is also important to be aware that lentiviral particles are a powerful...24h after transfection. This supernatant can be stored at 4 °C or can be directly added to your target ...

...including indels. Therefore, they engineered a dominant-negative mutant of the protein MLH1, a component of the...produced additive improvements. These variants are worth testing if you want to make your prime editing as efficient...Over 75,000 pathogenic genetic variants have been identified in humans and cataloged in the ClinVar database... potential to correct only a minority of those variants in most cell types. But prime editing, a CRISPR...ones in Addgene’s repository later in the post. Advantages of prime editing Less constrained by PAM sequence...research, the Liu lab created and evaluated 19 PE1 variants with RT mutations known to increase activity, ...came out on top? The Cas9 nickase fused to a pentamutant of M-MLV RT. They called this system PE2, which...

...distinguish between protein variants when used with a variant-specific antibody. TRIM21, an E3 ubiquitin ...sequence and RNA expression - but what if you just want to change protein levels? Until recently, no such...customizable protein degradation. Enter TRIM-Away - an antibody-based system developed by the James and Schuh ...recruits ubiquitin and the proteasome to destroy antibody-bound pathogens. Clift et al. found that addition...addition of exogenous TRIM21 protein along with an antibody targeting the protein of interest led to rapid...indicating that TRIM21 overexpression does not aberrantly activate the proteasome. Clift et al. tested ...co-microinjection of TRIM21 protein and the target antibody works well. In larger cell populations, you can...

...earlier this year: the first patient in the world, an infant, was successfully treated with a CRISPR gene editing... and mouse model systems, testing base editing variants, and assessing the safety of their intervention...disorder survive long enough to receive a liver transplant — creating intense time pressure for the research...double-stranded breaks in the DNA backbone, making them significantly less prone to indel mutations that plagued CRISPR...RNA-editing activity and developed a deaminase variant with a V106W mutation, which reduced RNA editing...editing efficiency. The V106W mutation was an important addition. Improved activity of adenine base editors... al., 2020).  Five years later, three of these variants (ABE8.8, ABE8.20, and ABE8e) would be tested for...