...cleavage promotes collateral cleavage of other RNAs. As with other Cas13s, this collateral cleavage does ...and Yan et al. describes new Cas13d enzymes that average only 2.8 kb in size and are easy to package in ...possibilities. To identify Cas13d, both groups used very general bioinformatic screens that looked for a CRISPR ...accessory protein RspWYL1 increases both targeted and collateral RNA degradation by RspCas13d. RspWYL1 also increased...

...to edit. These cells are then transfected with several different constructs for the reporter to work and...your CRISPR experiments. But how? There have been several systems to do this but most are labor-intensive... the native methionine start site. This first iteration was named GEmCherry1, due to the “genomic fragment...and 21 to create GEmCherry2. The Denham lab made several other small changes creating 2 other versions but...  Find GEmCherry2 reporter plasmids at Addgene Overall SRIRACHA and GEmCherry2 reporter systems provide...

...components of these vectors are also split across several plasmids (often three), for safety purposes. The...The 5’ LTR and 3’ LTR, respectively, act as RNA polymerase II promoter and terminator regions and are slightly...factors displayed on the outside of the particle to interact with the host. This is the role of Gag proteins...of lab evolution. There are 1st, 2nd, and 3rd generation plasmids for their production. As such, there...replication, with E2B being the primary viral polymerase. E4/E1A activate transcription of other relevant...

...previously a large barrier for gene therapy. In fact, AAV gene therapies have already been approved in the...non-integrating) as lenti and retro viruses do. They are generally used to achieve transient, high expression of ... Broad Tropism Only Infects Dividing Cells Moderate Immunogenicity Insertional Mutagenesis Possible...Lentivirus Listen to Connie Cepko Discuss Gene Therapy with AAV Figure out How to Get the Most from Your...

...and fostering team success. This shift requires deliberate preparation and a willingness to embrace new ...the room," understanding team motivations, and leveraging tools like the Myers-Briggs Type Indicator or...technical and managerial roles, and I've witnessed several colleagues do this while changing industries and...Courses: Leadership programs on platforms like Coursera, LinkedIn Learning, or edX. Podcasts: HBR IdeaCast...Scientists: What Makes a Good Leader Anyways?  Transferable Skills Guide: Managing a Team  ...

... a structural precursor protein, and Pol is a polymerase. Envelope gene (may be pseudotyped to alter infectivity...γ-retroviral packaging plasmids are not interchangeable. General envelope plasmids, such as VSV-G (Vesicular Stomatitis...packaging cell line. For example, Phoenix, a second generation γ-retrovirus packaging cell line developed by...region of the 3’ LTR and are primarily used in gene therapy research to reduce promoter interference and/or...More resources on addgene.org Addgene Protocols: General Transfection The Addgene Retrovirus Guide  ...

...single color controls On a flow cytometer, you generally want your positive cells to read between 104 and...provided by the antibody companies or check the literature. Pro tip! Early gates, like your live/dead markers...population (b). There is some debate in the literature as to when you need isotype controls. Our lab...isotype controls, while others don’t. However, the general consensus seems to be that if you are looking for...research background in CD8+ T cells and cancer immunotherapies. She is currently a research technician in ...

...the target molecule. Although it’s possible to generate antibodies against many types of molecules including...recognize proteins. Figure 1: Steps in the generation of hybridomas for monoclonal antibody production...19420862.2018.1445456  Holzlöhner P, Hanack K (2017) Generation of Murine Monoclonal Antibodies by Hybridoma ...(DC): National Academies Press (US); 1999. 1, Generation of Hybridomas: Permanent Cell Lines Secreting...hybridomas  Learn about monoclonal antibodies for immunotherapy Resource on Addgene.org Check out Addgene’...

...can I use it? CasPEDIA operates like many search engines, featuring both a general search bar tool and an... trans activity. AsCas12a, on the other hand, generates staggered DNA nicks at its target in cis and cleaves...resources throughout the site. In the Experimental Considerations section on each enzyme page, you'll find a ...packaging limitations. Maybe you want to read literature on the less common applications of an enzyme ...

...and grades that serve as automated filters of consideration. If you are worried about any eligibility provision... under two pages unless you are applying for a Federal Government opportunity, which tends to be more ...on grammatical errors, sentence structures, and overall format (fonts, font size, legibility, and more)...set of common questions with a peer or mentor. Several days prior to the interview, inspect your technology...come along for the ride, and consider applying to several opportunities. In an age of scarcity, the rational...

... software development team was working on lab operations software for next-gen sequencing (NGS), and high-throughput...did not. It was my job to translate complex lab operations workflows into a format the developers could ...could understand, so they could code the best operations software possible. Similarly, when the developers...a good fit for you if you are energized from interacting with others, and have strong communication skills... get software product management internships, preferably on teams that run within the agile framework ...

...publication in a scientific journal requires several considerations. First, it is important to establish the...whether your paper is in scope for the journal’s coverage. These guidelines also provide information on ...delay publication and manuscripts may go through several rounds of revision before acceptance. F1000Research...prerequisites of the journal. Ask colleagues (preferably native English speakers) to proofread your manuscript...

... cloning, you’re probably aware that there are several methodologies currently available for approaching... and Gateway® cloning technologies, as well as several others. Each method is unique and relies on specific...to propagate because CcdA wasn’t available to counteract the toxin. If, however, an investigator were ...answer lies in special strains of E. coli that tolerate the expression of the toxin gene. One such strain...plasmid CcdB protein involves poisoning of DNA-topoisomerase II complexes.” J Mol Biol. 1992 Aug 5;226(3)...

...studied a synaptic protein that is linked to neurodegeneration. The level of this protein is decreased in...method for the visualization of protein-protein interactions in situ. In the standard PLA assay, primary ...endogenous proteins of interests, suspected to interact, are added to a cellular sample. Both antibodies...visualization of the location of the protein-protein interaction. To study protein levels in situ, the Schumann...protein plasmids Use FRET to study protein-protein interactions Deposit plasmids with Addgene! ...

...on DNA purification without a kit that outlined several reasons why doing something without a kit has advantages...everything, including your skin. Here are some general tips for working with RNA, even if you are using...method, so biotechnology companies have marketed several products that help to further streamline this process...quantitative PCR to assess gene expression. There are several things that researchers can do to combat this issue...and proteins. Lastly, using a DNase (there are several DNase enzyme products on the market to choose from...

...repeating before you can finally dive deep into generating the data you’re excited about. When we were ...increase dramatically if you are piecing together several PCR products together. And this assumes you did...cost per stab has increased 50% since 2019, while overall inflation of USD has gone up 19.8% in the same ...making one plasmid at a time, try constructing several plasmids in parallel. That way, you can run one...help scientists save even more!) If you assume an average of 5 hours of hands-on time per plasmid, Addgene...

...means that F(ab’)2 fragments may have overall stronger interaction with their antigens than a Fab or Fab...the protein will interact with other components of your experiment. These considerations will help you narrow...secondary antibodies you already have in your refrigerator will not recognize a Fab fragment. In order ...

...secondary considerations to validation, application specificity, and experimental considerations - no other...rarely work for every assay. Antibodies that are generated against a linear antigen tend to work very well...permeabilization will influence how the antibody interacts with its target and consequently the quality of...isolate an antibody’s immune complexes and confirm interacting partners by mass spectrometry (Uhlen, 2016). ...

...genome editing in large-scale has led to the rapid generation of genome-wide gRNA libraries that knock out ...themselves incredibly useful for the efficient generation of individual (and multiple) genetic perturbations... al., 2015; Wong et al., 2016). CombiGEM uses iterative one-pot reactions with straightforward restriction...functionally characterize combinatorial gene knockouts generated via multiplexed gRNA expression (Wong et al., ...gRNA targeting sequences, you can then readily generate your barcoded gRNA library sequences via oligo...

...Collection at Coriell. More information and images generated from these cell lines can be found in the Cell...functional aspects of the protein gleaned from the literature and engagement with researchers who study the...find out more about gene edited hiPS cell lines generated using the Allen Institute for Cell Science Plasmid...Drosophila cells, and C. elegans. In addition to generating fluorescent protein fusions, plasmids are available...find out more about gene edited hiPS cell lines generated using these plasmids, please visit the Cell Catalog...