... gRNAs By Species Mammalian Bacteria Drosophila Plant C. elegans Yeast Zebrafish Xenopus CRISPR Resources...target the dCas9-activator to your specific locus. Want more information on the wide variety of Cas enzymes...expression in mammalian systems, bacteria, Drosophila, plants, and yeast. Mammalian ID Plasmid Gene/Insert Promoter... ID Plasmid Gene/Insert Promoter PI Publication Plant ID Plasmid Gene/Insert Promoter Selectable Marker...

... gRNAs By Species Mammalian Bacteria Drosophila Plant C. elegans Yeast Zebrafish Xenopus CRISPR Resources...the collateral RNA degradation seen in bacteria. Want more information on the wide variety of Cas enzymes... expression in mammalian systems, bacteria, and plants. Mammalian ID Plasmid Gene/Insert Promoter Selectable...Bacterial ID Plasmid Gene/Insert PI Publication Plant ID Plasmid Gene/Insert PI Publication Do you have...

... gRNAs By Species Mammalian Bacteria Drosophila Plant C. elegans Yeast Zebrafish Xenopus CRISPR Resources...HDR occurs at a much lower efficiency than NHEJ. Want more information on the wide variety of Cas enzymes...expression in mammalian systems, bacteria, Drosophila, plants, C. elegans, yeast, zebrafish, and Xenopus. Mammalian...Insert Promoter Selectable Marker PI Publication Plant ID Plasmid Gene/Insert Promoter Selectable Marker...

... gRNAs By Species Mammalian Bacteria Drosophila Plant C. elegans Yeast Zebrafish Xenopus CRISPR Resources... increases off-target effect. Which species or variant of Cas9 ( S. pyogenes , S. aureus etc.) was this...compatible with a single species or PAM binding variant of Cas9. For instance, wild-type SpCas9 must be...information and assistance with the deposit process. Want more information on the wide variety of Cas enzymes...

...mild to severe respiratory disease in humans. Advantages of adenoviruses in research include their large...adenoviruses do not integrate into the host genome. Disadvantages of adenoviruses include transient transgene ...transgene expression, as well as the substantial immune response induced by the viral capsid. Adenoviruses can...Plasmids Common shuttle and adenoviral plasmids. Want an adenoviral plasmid with your gene of interest...

...protein, and is suitable for both cell-permeant and non-cell-permeant proteins. CLIP-tag CLIP-tag is identical...proteins with disulfide bridges such as antibodies. MCP-tag is a mutant of ACP-tag, and is labeled by SFP Synthase...

...Loss of the tumor suppressor p53 also promotes aberrant mTORC1 activity, and multiple familial cancers...uncontrolled cell growth and proliferation, including an abundant supply of macromolecules and energy, as well as... as the presence of both pro-proliferative and anti-apoptotic signals. mTORC2, which appears to signal... mTOR signaling in growth control and disease. Laplante M, Sabatini DM. Cell. 2012 Apr 13;149(2):274-93...

...common antibiotic resistance genes and which applications are best (or worst) for each antibiotic. The... as SHuffle cells, are already resistant to this antibiotic due to stable integration of resistance genes...-in-one antibiotic — it is usable in eukaryotic cell culture, yeast, bacteria, and even plants! The zeocin...host DNA damage. Choosing an antibiotic resistance Choosing the best antibiotic resistance ultimately comes...pay attention to antibiotic selection. If you feel like multiple of the above antibiotics would work for... the pros and cons of each antibiotic and happy selecting!   Antibiotic Resistance Stability Time ...Over-Agar Antibiotic Plating Plasmids 101 eBook Resources on the Addgene blog Plasmids 101: Antibiotic Resistance...

... blog Antibodies 101:Selecting the Right AntibodyAntibodies 101: Secondary AntibodiesAntibodies 101: Buffers...Addgene.org Addgene's Antibody GuideAddgene's collection of ready-to-use recombinant antibodies...a given experiment, it is important to consider how many parameters you want to use in a given experiment...detector using a series of mirrors and filters. Importantly, conventional flow cytometry uses one detector...conventional and spectral flow cytometry, it is important to consider the expression levels of the markers...two highly overlapping cellular markers, it is important to pair them with two fluorophores that have distinct...is critical that your controls and your sample antibodies are from the same batch, to avoid unmixing issues...

...ahead of time that you want to probe your blot with multiple antibodies. You’ll want to set up your blot ...starting with the least-abundant protein and working toward the most abundant. If the abundance of your...isn’t too different, consider antibody affinity: stronger binding antibodies will require harsher stripping...without a re-wetting step. Important: do not dry your membrane with antibodies on it if you plan to reprobe... avoid retaining residual antibodies. If you have already applied antibodies and need to dry the membrane...stripping solution uses low pH to disrupt antibody binding to antigens. Ingredient Concentration or ...you don’t see any signal, you’ll want to check for the primary antibody by re-blocking your membrane and...

...Addgene blog Antibodies 101: Flow Cytometry  Antibodies 101: Reading a Flow Plot Antibodies 101: Conventional...case through conjugated antibodies targeting the cell surface markers CD11 (anti-mouse CD11b-BV510), which...looser gate here. I am not worried about including irrelevant events in my analysis, as these will get excluded...Gating Having gated my single lymphocyte cells, I now want to exclude all dead cells from my analysis. For ... protect themselves from it. So here, I actually want to gate on my negative, unstained population, which...experimentally-specific gates. Two-dimensional staining (quadrant gating) For our last example plot, I will show...present on dendritic cells but not T cells, and CD8 (anti-mouse CD8b-PerCP-Cy5.5), which is present on cytotoxic...

...Reactions Anti-drug antibodies (ADAs) are formed when the body recognizes a drug as a foreign antigen, triggering...replicate the complexity of antigen processing events that take place within an antigen-presenting cell like ...Beijnen, J. H., & Schellens, J. H. (2016). Antidrug Antibody Formation in Oncology: Clinical Relevance .../s12929-019-0592-z  Resources Antibodies 101: Introduction to Antibodies CRISPR in the Clinic   ...Monoclonal antibody drugs are popular therapeutics for a plethora of disease conditions, from cancer ...cancer to autoimmune disorders. Antibodies administered as drugs are still immunogenic, meaning that they elicit...of immunogenicity can help evaluate risk of an antibody.  In silico prediction of adverse reactions In...

...bacteria, it is important to note that the term ‘antibiotic’ can also be used to describe anti-parasitic drugs... is not naturally resistant but has become resistant somehow overtime. One fantastic example of intrinsic... spread of antibiotic resistant bacteria, check out this resource on preventing antibiotic resistance ...Addgene's Microbiology Pages What is antibiotic resistance? Antibiotic resistance is defined as microbial... to antibiotics designed to kill them or to impede their growth. When we hear the word ‘antibiotic’, very... drugs and anti-fungal drugs, because ‘antibiotic’ merely means that the drug prevents the growth of a...treat. Learn How Antibiotic Resistance Genes Are Used in the Lab How does antibiotic resistance develop...

...this logic, ω-dCas9 variants recognizing multiple PAMs should have a fitness advantage over proteins with...flat out better than SpCas9. I want everyone to test it because I want to know the answer", says Liu. ...work, there are other options. Commonly used Cpf1 variants, with a  5’ TTTV PAM, improve targeting in AT-...respectively. Researchers also engineered SpCas9 variants with alternative PAMs like NGAG and NGCG and SaCas9...phage-assisted evolution to generate and select for Cas9 mutants with PAM flexibility. In this system, the mutagenesis...promoting survival of only the broadest-targeting variants. For the final round of evolution, they used a... a no-G PAM library to enrich for Cas9 variants with activity on non-NGG PAMs, generating xCas9 3.0-3.13...

...expression of antibodies in plants using syringe agroinfiltration." Recombinant proteins from plants. Humana...transgenic plant can take even longer, a transient plant leaf transformation process could save the plant biologist...Browse our Plant Plasmids and Resources page Find synthetic biology plant plasmids Find CRISPR plant plasmids...vincristine, are regulated in the medicinal plant, the Catharanthus roseus (Figure 1). The ultimate goal of...to enhance their production either in the plant or in plant tissue cultures.   C. roseus is not a model...pathway. Why plant scientists do transient leaf transformations The development of transgenic plants is a common...the plant sciences “transient transformation” usual means that neither a stable cell line or plant is ...

...improve your grant writing skills. Good luck! Further advice on grant writing Since successful grant writing...with your work, your advisor may want to mentor you during the grant application process. When my PhD ...have no idea how to write a grant!”, “I don’t have time for this if he wants data by next week!”, and “...RFA contains all of the important information required for a successful grant application. It typically... advice on writing a grant proposal. "Murder Most Foul: How Not to Kill a Grant Application" by Vid Mohan-Ram...How to write a research project grant application" by the NIH Still want more information on the application...their advisor alone applies for these competitive grant applications; however, in some cases, your advisor...

...GreenGate – offers all of those same advantages to researchers working in plant model systems. “The cloning is... most frequently used elements in plant expression cassettes: plant promoters, N-terminal tags, coding...C-terminal tags, plant terminators and resistance cassettes for selection of transgenic plants. The complete... in vitro; it’s no different in mouse, human or plants,” explained Jan Lohmann of Heidelberg University...based on the daily needs of an advanced molecular plant science lab.” Introducing GreenGate As Lohmann ...required to generate multi-construct transgenic plants. In the most practical terms, what GreenGate can...Arabidopsis, it should in principle work in any plant species. Tricks of the trade Lohmann does have one...

...single-stranded breaks ( nicks ). ID Name Insert Function Antibiotic Resistance Activity PI 52962 lentiCas9-Blast ... a transcriptional activator . ID Name Insert Antibiotic Resistance Activity PI 61422 dCAS9-VP64_GFP dCAS9... express fluorescent proteins. ID Name Insert Antibiotic Resistance Purpose PI 17446 pLenti CMV GFP Hygro... in a new window) . Please note this does not guarantee viral service, but lets us know what viruses would...

...for use with virus-mediated delivery has many advantages for research. Viruses facilitate the delivery...Stable RNA <8 Kb 80-130 Dividing Yes Moderate * Recombinant AAV has a low frequency of target cell genome... Lab Lentivector Toolbox Guide to Generating Recombinant Adenoviruses Eric Campeau's Viral Vector Guide...Biosafety Resource Guide . References Cohen S, Au S, Panté N. How viruses access the nucleus. Biochim Biophys...

...during Development and Regeneration - Periklis Pantazis Lab Multibow: digital spectral barcodes for cell... loss and ablation of previously resistant cell types. CreLite - George Eisenhoffer Lab. ...for Zebrafish Receptors and Zebrafish Monoclonal Antibody plasmids - Gavin Wright Lab. Collections of plasmids...that encode zebrafish receptors or monoclonal antibodies to zebrafish cell surface receptors and secreted...