...made smaller. Previous attempts to “shrink” Cas9 include the use of St1Cas9 (~3.3 kb) from Streptococcus... approximately 1350 amino acids in size, which includes SpCas9, and shortern orthologs approximately 1000...in mouse liver.  Two other smaller Cas proteins include Cas12b and CasX. While there are no papers yet ...capabilities of full-length SpCas9. This set of plasmids includes AAV-Cas9C-VPR for targeted gene activation. Split...

...but some examples I’m personally interested in include: intracranial self-stimulation (ICSS) and place...resolve these issues is with electrophysiology. Concluding thoughts and moving beyond the mouse While it...applicable to the rat [6]. Technical differences include the ferrule implants and patch cables. One huge... at the Rockefeller University.  His interests include the role of learning and memory in opioid addiction...

...depend on target and sample type. Example steps include: permeabilization, antigen retrieval, and sectioning...available, and your experimental goals. Example steps include: counterstaining, mounting sample on slides, and...you optimize your protocol, always remember to include controls to show that your antibodies are binding...antibodies validate them in a variety of applications, including IF (or IHC or ICC), and will share some of their...

...Several factors contribute to the popularity of GFP including (i) fast and complete maturation to functional...endogenous fluorescent co-factors can be used. Examples include the LOV domains that bind flavin (Buckley et al... conditions, with pKa values below 4. Examples include mTurquoise2 (pKa = 3.1), tagRFP (pKa = 3.1) and... or media. These sources of ‘autofluorescence’ include riboflavin in the medium or flavin containing proteins...

...AAVs to broadly spread throughout an organism, including difficult-to-target tissues like the central nervous...and so on, although we often use abbreviated nomenclature (AAV1, AAV2, etc.) to distinguish the capsids...AAV pseudotyping. It has broad tissue tropism, including liver, muscle, lung, CNS, kidney, and retinal ...serotypes used to target the central nervous system, including neurons, AAV5 is generally the best choice for...

...for CRISPR genotyping. The most common options include mismatch cleavage assays, such as Surveyor™, T7E1...information profile regarding putative off-target sites including both the edit rate and the repair product sequence...algorithms do not take into account other elements, including those relating to DNA structure (e.g. epigenetic...options exist for unbiased off-target detection, including Digenome-seq (Kim et al. 2015) for in vitro analysis...

...afflicted by pervasive off-target effects. This conclusion was based upon numerous phenotype discrepancies...importance of using orthogonal methods to confidently conclude the function of a gene of interest. Tools for ...discussed below). Examples of these constructs include pGAPi, which allows easy creation of long hpRNA...experiment are now available on Addgene. These include the construct that allows for easy insertion of...

...various sources that are difficult to trace. These include the laboratory personnel, the serum, the cell culture...cross-contamination and due to poor lab techniques. These include reusing pipettes for multiple cell lines rather...Contamination! Tissue culture media often have indicators included in the media that turns yellow from lowered pH... periods and affect the total cell viability. Conclusion To sum up, it is always important to prevent ...

...organizations in a variety of science policy roles including fellowships, volunteer positions, and part-time... with a large set of other societal pressures including economic, cultural, and political. Science policy...science blogs. Other avenues for skill development include leadership of extracurricular organizations, working...opportunities. NSPN maintains a great resource guide including conferences, societies, and outreach groups. Grab...

... molecule at all, leading to inconclusive results, erroneous conclusions, and wasted time. In fact, many...antibody-dependent approach. Antibody-independent methods may include analysis of transcriptomic or proteomic data from...specifically to the expected protein. Common approaches include using RNAi to knock-down expression of your target...

..., introduced the “Three Es of Immunoediting” nomenclature. 1. Eliminate - Latent tumors are recognized...on immune system Conventional cancer therapies include radiation therapy (owing to contributions from ...blocking. Other molecular targets under research include TIM-3, LAG-3, STAT-3, EGFR, CD20 and CD73 on specific... factors researchers are beginning to focus on include: Cell heterogeneity Tumors are composed of a wide...

... Figure 1: Labeled diagram of an antibody including Fc, Fab, heavy chain, light chain, constant region...antibodies for experiments in a variety of fields including neuroscience and immunology. They can be used ...analyzed using techniques such as flow cytometry. Conclusion Whew! Now you know that antibodies can be used...used for. Addgene’s Antibody Plasmid Collection includes plasmids that express antibodies, nanobodies, ...

...for plasmids suitable for other model systems, including E. coli, plants, Drosophila, and zebrafish!   ...destination vectors. Destination vector options include humanized wt Cas9, dCas9 (transcriptional repressor... Figure 3: Comparison of Multiplex Strategies including Standard PolIII-gRNA cassette, Csy4-cleavable ...filler sequences. You also have the option of including a previously-designed tyr gRNA, which causes hypopigmentation...

...for plasmid. pXXXXX-XXXX Step 1: Backbone name Include the empty backbone name in your plasmid name. This...backbones, visit Vector Database. Step 2: Insert name  Include information about the insert in your plasmid name...contains a mutation or modification, this should be included in the plasmid name. Mutations are generally listed...

...species, antibody species, or whether the results include knockout data.    Figure 2: Search options in ...description of the experiment. If that data was included in a publication, you'll find that information... is easy!  Here at Addgene, we love all data, including negative data, which can help researchers save...

...the template. The PE3 system does just this by including an additional sgRNA. Using this sgRNA, the prime... number of unintended prime editing outcomes, including indels. Therefore, they engineered a dominant-...     ✓ ✓ reflects an innovation always included in the indicated tool. ~ reflects an innovation... strategies in our CRISPR guide. Some examples include: TwinPE: two prime editors make complementary ...

...per gene and includes ~10,000 gRNAs targeting intergenic regions. The libraries also include 2,400 gRNAs...recently deposited to Addgene and you’d like it to be included in the next What’s New in CRISPR blog post, please...

... to a post-academic career. This training was “included” with their tuition, so to speak.  Unfortunately...the development of priorities for pd|hub which include building capacity to identify the most effective...scientists, but it is essential that we integrate inclusive career readiness training in this educational ...

...White House honored 13 “Champions of Change,” including our recent guest Drew Endy, in celebration of ...trouble, even when all kinds of other factors, including journal impact factor, study topic, and the corresponding...review. 8)     That same issue of Science also included an online survey of scientists showing overwhelming...

...attracted much attention over the past few years, including an article in the BBC Wildlife Magazine. It is...people for years and have gathered much acclaim - including being featured in the National Geographic and ... are now taking place and gathering momentum, including in Cambridge and Bristol. If you’re a scientist...