Skip to main content
Addgene

Enter a sequence to perform a BLAST-based search of the Addgene plasmid sequence database.

Information

In Category

We narrowed to 884 results for: ANT;

Showing: 181 - 210 of 884 results

  1. Meet an Addgenie: Insights into Grad School, Career Paths, and More

    Type
    Blog Post
    Published
    June 25, 2020, 1:15 p.m.
    ... 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...

  2. Progress Towards a PAM-Free CRISPR

    Type
    Blog Post
    Published
    March 7, 2024, 2:39 p.m.
    ...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 ...

  3. AAV Vector Quality Control: Going the Extra Mile with NGS

    Type
    Blog Post
    Published
    Sept. 12, 2017, 1:44 p.m.
    ...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%. ...

  4. Finding Your Perfect Job After University

    Type
    Blog Post
    Published
    Jan. 12, 2016, 3:30 p.m.
    ... 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...

  5. Multifunctional Neural Fibers: Modulation from Gut to Brain

    Type
    Blog Post
    Published
    Aug. 31, 2023, 1 p.m.
    ... 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...

  6. Plasmids 101: Protein tags

    Type
    Blog Post
    Published
    Dec. 11, 2014, 4:26 p.m.
    ...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 ...

  7. Advancing Biology with Zebrafish: Genetic Tools for Developmental Studies and More

    Type
    Blog Post
    Published
    Aug. 13, 2019, 12:58 p.m.
    ...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...

  8. What's Your Organism? Expanding Genomic Tools via the NSF EDGE Program

    Type
    Blog Post
    Published
    May 2, 2019, 12:41 p.m.
    ...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....

  9. Four Base Editing Reporters to Monitor and Enrich Editing in Real-time

    Type
    Blog Post
    Published
    July 7, 2020, 1:15 p.m.
    ...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...

  10. CRISPR 101: Which Cas9 Do I Choose for My CRISPR Experiment?

    Type
    Blog Post
    Published
    Oct. 13, 2022, 1:45 p.m.
    ...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...

  11. Quick Guide to All Things Lentivirus

    Type
    Blog Post
    Published
    March 21, 2017, 2:30 p.m.
    ...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 ...

  12. Prime Editing: Adding Precision and Flexibility to CRISPR Editing

    Type
    Blog Post
    Published
    Jan. 13, 2025, 9:49 p.m.
    ...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...

  13. Hot Plasmids April 2018 - Protein Degradation, Nanoscopy, FIRE-Cas9, and Yeast Expression Tools

    Type
    Blog Post
    Published
    April 23, 2018, 2 p.m.
    ...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...

  14. Proximity Labeling: A Powerful Tool for Protein Complex Purification and Proteomic Mapping

    Type
    Blog Post
    Published
    Dec. 2, 2019, 2 p.m.
    ...directed evolution until they identified a mutant with significantly increased sensitivity that did not affect...compared to the standard E. coli BirA* mutant. RapID consists of this mutant BirA* and an RNA component that ...elucidate these interactions scientists have long used antibody based affinity purification and mass spectrometry...proteins, which can be a challenge for classic antibody based affinity purification/mass spectrometry ...proteins resulting in their biotinylation. The advantage of APEX is its speed, labeling neighboring proteins...calcium uniporter (Martell et al., 2012). It is important to note that the biotin-phenol reagent can be ...original APEX (Lam et al. 2014). They named this mutant APEX2.  Find the APEX2 plasmids! TurboID and minturbo...

  15. Site Directed Mutagenesis by PCR

    Type
    Blog Post
    Published
    Aug. 2, 2016, 2:30 p.m.
    ...plasmid context). The technique is also highly relevant in this age of CRISPR; site-directed mutagenesis...facilitate the repair, the template should be resistant to Cas9 cleavage. If a plasmid contains the template...cleavage), thereby rendering the resulting construct resistant to Cas9 induced cleavage. Summary of Site Directed...and bacteria are transformed with the nuclease-resistant nicked plasmid (the PCR product). Plasmids are...interest. Gel electrophoresis diagrams of the anticipated restriction analysis results (left) and validation...figure). Template A high purity plasmid prep significantly increases the success rate of site directed ...directed mutagenesis. In addition, you may want to try different concentrations of template (e.g. 0.1-1.0 ng/...

  16. Addgene Launches Partnership with IPI

    Type
    Blog Post
    Published
    June 13, 2023, 1:15 p.m.
    ... ready-to-use recombinant antibodies. The first collection, a set of integrin antibodies, will be available...website and blog, including our antibody guide, antibody protocols, and Antibodies 101 blog series.  Stay tuned...s website this fall. IPI antibodies will come with access to the antibody sequences, in line with the...the rest of Addgene’s recombinant antibody collection, as well as access to IPI educational resources ...resources developed to support use of their antibody tools. Addgene also offers a growing collection of educational...

  17. Plasmids 101: Positive and Negative Selection for Plasmid Cloning

    Type
    Blog Post
    Published
    Aug. 22, 2019, 12:43 p.m.
    ...includes an antibiotic resistance cassette and you aren’t introducing an additional antibiotic resistance...selection strategy, here are a few to consider. Antibiotic selection Selection is straightforward in cases...cases where the inserted DNA includes an antibiotic resistance marker. After transformation, plate the ... cells on medium containing the corresponding antibiotic. For example, if the DNA you’re inserting into...along with your gene of interest. In these cases, antibiotic selection will select for cells that have been...contains your gene of interest. Auxotrophy Besides antibiotic selection, you can select for a successful uptake...uptake of your DNA of interest by taking advantage of bacterial characteristics such as prototrophy, the...

  18. Plasmids 101: What is a plasmid?

    Type
    Blog Post
    Published
    April 2, 2020, 6:17 p.m.
    ...typically include a gene that is advantageous for survival, such as an antibiotic resistance gene. In contrast...expressed in and amount of recombinant protein obtained. Selectable Marker The antibiotic resistance gene allows...bacterial cells and selectively grown on antibiotic plates.  Importantly, because the bacteria from which plasmids...components that make up a plasmid and why each is important. Our “Plasmids 101” series designed to educate...naturally in archaea and eukaryotes such as yeast and plants. In nature, plasmids provide one or more functional...functional benefits to the host such as resistance to antibiotics, degradative functions, and/or virulence. All...by recruiting replication machinery proteins Antibiotic Resistance Gene Allows for selection of plasmid-containing...

  19. Production of Virus in Insect Versus Mammalian Cells

    Type
    Blog Post
    Published
    Nov. 5, 2024, 8:32 p.m.
    ...straightforward — simply spin down the supernatant in a centrifuge to remove any unwanted cellular debris, as you would...produced in multiple different cell types, including plants, mammals, and insects. In this post, we will compare...which we talk about below. Processing The viral supernatant is typically collected over the course of a couple...to 24 hours. Processing the virus-containing supernatant will depend on the virus type. AAV is typically...methods like ultracentrifugation, while lentivirus supernatants are centrifuged and filtered to eliminate any...than a P1 virus. Pro-tip! Centrifuge the viral supernatant in between passages to eliminate dead cells and...of production, saving about a week of work. An important note is that this is only possible because baculoviruses...

  20. CRISPR 101: Off-Target Effects

    Type
    Blog Post
    Published
    Feb. 8, 2024, 2:15 p.m.
    ...several options to quantify these events. Whole genome sequencing The only way to quantify off-target effects...there are significant editing events, serving as a proxy for total off-target effects. Quantifying off-targets...CRISPR work, this often means DNA cleavage at unanticipated sites. While off-target effects are nearly impossible...and minimize them, and ways to control for and quantify them when they can’t be avoided. What are off-target... to keep in mind, but likely will not have a significant impact on your data. You are trying to generate...better to reduce off-target effects, but it’s important to not let ‘perfection’ get in the way of ‘good...mispairing of bases. To reduce off-targets, you’ll want to select a Cas enzyme with low tolerance for mismatches...

  21. Genome-wide Screening Using CRISPR

    Type
    Blog Post
    Published
    Aug. 18, 2015, 2:30 p.m.
    ...example, mutant cells can be used in drug screens to identify genes that confer drug-resistance. Mutant cells...library plus Cas9 to generate the mutant cells, respectively (C). Mutant cells are screened (D) and "hits...post was updated on Aug 20, 2020. What genes are important in your phenotype of interest? Many scientists...not entirely known. Identifying which genes are important for a phenotype can lead to a wealth of additional...Addgene.  What makes CRISPR so special? A major advantage of CRISPR/Cas9 over previous genome editing techniques...experiment is to generate and screen a population of mutant cells to identify genes involved in a particular...library to generate a heterogeneous population of mutant cells, with each cell or set of cells containing...

  22. Deciphering the Mysteries of Behavior with Viral Vectors

    Type
    Blog Post
    Published
    Sept. 17, 2019, 12:40 p.m.
    ...unintentionally remove connecting pathways between other distant regions. They also don’t give any clues about the...and depression.  Locomotion tests To study and quantify locomotion, scientists monitor an animal’s movement...rather than being out in the open (risky). Taking advantage of these characteristic traits, rodents can be...used since the 1970s to test the efficacy of antidepressant treatment in rodents. In this test, an animal...BDNF in the hippocampus causes a shift towards antidepressant behavior in rats in the forced swim test as...neurons (Drd1) in mPFC is responsible for the antidepressant effect. To specifically activate the Drd1 neurons... of microRNA124a in the hippocampus evokes antidepressants-like effects in rats." Psychoneuroendocrinology...

  23. Hot Plasmids - August 2020

    Type
    Blog Post
    Published
    Aug. 18, 2020, 1:15 p.m.
    ...PAC-MAN (prophylactic antiviral CRISPR in human cells) is a CRISPR-Cas13 based antiviral method to degrade...protease used to cleave affinity tags from purified recombinant proteins at small and large scale.  This new ...utility of the system for use with low affinity variants of TEV. The new system has been demonstrated to...the evolution platform to generate TEV protease variants. Catalysis by TEV protease allows a transcription... The benefits of nanobodies over conventional antibodies can’t be overlooked. Nanobodies are smaller in...easier to clone, express, and purify compared to antibodies. Nanobodies are easily expressed in the cytosol...of bacteria where it can be secreted into the supernatant. Four plasmids containing different nanobodies...

  24. Fluorescent Proteins 101: Photoactivatable Fluorescent Proteins

    Type
    Blog Post
    Published
    April 25, 2017, 2:30 p.m.
    ...designed 2 variants of EosFP that can withstand these conditions- mEos4a and mEos4b (12). Both variants are ...light (~400 nm), the chromophore photoconverts predominantly to the anionic form, causing increased fluorescence... random mutagenesis screening for enhanced RFP variants. PA-mCherry (E26V / A58T / K69N / L84F / N99K ...respectively. When illuminated with violet light, both variants are photoconverted to forms that emit brighter...PA-FPs. Several other irreversible PA-FP spectral variants have been discovered such as Kaede and PS-CFP2...localization, or leading to protein aggregation. Another important parameter to consider is the contrast ratio- a... are performing multicolor experiments, you will want to be sure that the light used to activate the first...

  25. Harnessing Bacterial Toxins for Allelic Exchange

    Type
    Blog Post
    Published
    Aug. 15, 2019, 12:30 p.m.
    ...be replicated in the recipient, the only antibiotic resistant colonies isolated are those where the allelic...single crossovers, they are just spontaneous antibiotic resistant clones. But if they are blue or magenta,...hospital-associated Gram-negative bacteria become resistant to antibiotics! References Espah Borujeni, Amin, Anirudh...chromosome is to disrupt expression of a gene with an antibiotic resistance cassette. This leaves a “scar” in ... This can be accomplished by selecting for an antibiotic resistance marker from the vector (abR, Figure...efficient method to generate precise, unmarked mutants, but it frequently requires considerable optimization...mix of Type VI secreted effector toxins, toxin-antitoxin type toxins, and phage holins. They hypothesized...

  26. Transferable Skills Guide: Conflict Resolution

    Type
    Blog Post
    Published
    Jan. 4, 2022, 2:15 p.m.
    ...our Transferable Skills series, this post is not meant to be a comprehensive guide to Conflict Resolution...explaining my approach to conflict resolution, it’s important to recognize that there are different types of... poor decision making, and stalled projects. You want to identify and address these types of harmful conflict...conference poster session, you don’t necessarily want everyone who visits your poster to agree with your...also be categorized as either “personal” or “substantive”. A personality-based conflict occurs when two...a lab or work environment if left unchecked. Substantive conflict occurs when people or teams who normally...or goal. Resolving these types of conflicts is important for the individuals involved, and also for the...

  27. New Optogenetic Tools for Cytoskeleton and Membrane Control

    Type
    Blog Post
    Published
    Sept. 28, 2023, 1:15 p.m.
    ...signaling — think of a mouse with an LED brain implant or a worm wiggling back and forth under blue light...microtubule-cutting sword Microtubules are one of the most important components of the cytoskeleton. They provide cells...disrupt microtubules in cells, they have various disadvantages and limitations – for example, microtubule-destabilizing...are also available as lentiviral plasmids if you want to make a stable cell line. This design allows highly...that means this light-powered sword truly is an elegant tool. The authors demonstrated its use to probe... what if you’re interested in that other most-important component of the cytoskeleton – actin? Actin dynamics...on BcLOV4. These single-component systems take advantage of BcLOV4’s light-induced binding of the plasma...

  28. Tips for Getting a Faculty Position

    Type
    Blog Post
    Published
    July 20, 2023, 1:15 p.m.
    ... to Get a Faculty Position. This edition has substantial expansions on the chalk talks and Zoom interviews...emphatically deny that I have a magic formula to guarantee a reader the faculty job of his/her dreams. There...currency of scientific success is publications and grants. 2. Write succinct and easy to understand documents..., documents need to minimize jargon. Equally importantly, you really are selling yourself. A sales pitch...years. You will discuss the focus of your first grant, how you will overcome certain obstacles, alternative...not recommend putting these into slide format. You want people to focus on you and your ideas. Slides will...anyone could relate to, that people would agree is important to understand. (1-2 sentences). For example: Distinguishing...

  29. Thoughts on the Future of Research 2015 Boston Symposium

    Type
    Blog Post
    Published
    Nov. 25, 2015, 4 p.m.
    ...paid on fellowships instead of grants such that PIs no longer save grant money by paying relatively low...discussed during the symposium will prove incredibly important to the stability of the biological research enterprise...organizers indicate that the main question they wanted to focus on during the symposium was “How many ...some of the more detailed questions the organizers wanted to touch on in organizer David Riglar’s blog post...solutions. Keynote speaker Paula Stephan did a fantastic job of articulating many of these problems and...Stephan’s Talk (All slides can be found here). Importantly, the panel offered some great solutions to these... Talk to Your Editors I wish I had been more cognizant of this during my PhD. Submitting a publication...

  30. Troubleshooting Your Plasmid Cloning Experiment

    Type
    Blog Post
    Published
    Sept. 24, 2019, 12:55 p.m.
    ...colonies, check that the antibiotic in the plate correspond to the antibiotic resistance marker present...hopefully obtain your coveted plasmid with no substantial delays. Design, design, design Before diving ...fragments. You can generate the parts of DNA that you want to assemble together in a combination of different...difficulty amplifying your target sequence, or if you want to amplify sequences from a large genome organisms...successful and ultimately faster cloning. Remember to quantify the DNA concentration of all the samples by gel... experimental design. Full lawn of cells. The antibiotic in your plate might not work. Check the plates...E. coli  strain known to be sensitive to that antibiotic, and if the strain is growing fine, it is probably...
Showing: 181 - 210 of 884 results