We narrowed to 965 results for: REP

... homology-directed repair (HDR) to correct double-strand breaks (DSBs). DSB repair by HDR facilitates ... a large cohort of genes (e.g. tagging genes representing a protein super family). To overcome these challenges...complexed with sgRNA. Direct comparison of Cas9 RNPs prepared with an in vitro-transcribed (IVT) sgRNA versus...without investing time and labour on reagent preparation in the lab. Figure 3: csRNP-assisted epitope...mutations (e.g. convert the NGG PAM to NGC in your repair template) and use homology arms of 60 -70 bp . ...temperature for 10 min. Deliver csRNP into cells: Prepare cells for transfection, add csRNP and ssODN to ...

...generate dsRNA as part of their infection and/or replication process within the host. dsRNA viruses are inherently...generate dsRNA intermediates as part of the viral replication process. dsDNA viruses yield dsRNA through a ...RNA that the RNAi is designed against), RISC can repress translation and/or degrade target mRNA. These ... by bidirectional transcription or as inverted repeats. These elements are almost always epigenetically...particularly enriched in highly transcribed regions and repetitive regions, such as telomeres and centromeres. These...presumably due to their abundance of R-loops and repetitive sequence. All of the R-loops described above...

...line./ Each color represents a gRNA targeting a specific chromosome. This image represents the composite ...(GAL4) was supplied. In addition to labeling repetitive telomeric sequences, Chen et al. successfully...distance! To the authors’ knowledge, this work represents the first mapping of intrachromosomal loci, a...previously described for transcriptional activation/repression and genome editing. Ready to make your cells...engineering for genome editing, activation and repression.” Nat Methods. 12(11) (2015):1051-4. PubMed PMID...

...propagated it, separated it into roughly identical replicate groups, and treated these groups with the same...below): 1. Resistant cells among the separate replicates would contain the same labels - i.e. they would...would always come from the same parent cells that represent the preexisting resistant subpopulation 2. Resistant...Resistant cells among the separate replicates would have different labels - i.e. they would stochastically... Looking at barcode abundances among multiple replicate populations, they found that similar barcodes ...populations were shared by at least one other replicate; prior to treatment, there are small (previously...

...GCaMP6 family of sensors are commonly used calcium reporters with different kinetic parameters. There are three...also available. Detection methods Many calcium reporters use a single fluorescent protein (FP), requiring...unwanted variables. Twitch is a FRET-based calcium reporter derived from troponin C. It is engineered with...for in vivo studies) There are several calcium reporters specially designed for the aims and requirements...the target cells, ensuring more consistent and reproducible results in calcium assays. There’s another crucial...to go. Pro tip: Vectors with ready-to-use AAV preparations are labeled with the green virus particle on...

...New to the bench? Fragmid depositor John Doench reports even the greenest members of his lab, high school...plasmid creation process. Each module of Fragmid represents a class of components, such as your CRISPR enzyme...modular cloning approach, it’s easy to switch out reporters, Cas proteins, guide cassettes, promoters, N’ ...ampicillin resistance cassette and an origin of replication, which are not schematized. Sizes of fragments...always, we recommend checking your cloning via a miniprep and restriction digest, and then plasmid sequencing...skip a module entirely, but instead will have to replace it with a spacer.) Each component available on ...

...specificity.  Strategy #3 (ProC): Create promoters by repeating mouse cell type specific transcription factor ...expression of the same transgene: an optogenetic reporter, channelrhodopsin, fused to a fluorescent GFP ...expression. Surprisingly, <1% of synthetic promoters replicated the expression specificity of the wild-type promoters...database of the microscopy images of all AAVs that reproducibly labeled retinal cells can be found here.  Applications...expression of a Cre-dependent fluorescent CatCH-GFP reporter. Expression of GFP will only result in cells where... Find AAV plasmids Find ready-to-use AAV viral preps Browse all viral vectors References Juettner, J...

...able to facilitate and enhance DNA repair via homology directed repair (HDR). This characteristic makes ...potentially be treated by correcting the mutation or replacing the gene. A notable example is the treatment of...reconstitution with a normally functioning T cell repertoire (1). In this example, it was possible to transduce...first discovered as a contaminant of adenovirus preparations. It is an icosahedral, non-enveloped virus that...enhances endonuclease delivery for double-strand break repair." Gene therapy 17.9 (2010): 1175-1180. PubMed PMID...

...as an NPA representative and speak the NPA’s stance. I had to be professional as I represented the organization...As the Chair of the Advocacy Committee, I had to report to the Board of Directors and liaise with the Executive...accordingly. For example, the quarterly and annual reports I had to write on behalf of the Advocacy Committee...to overcome the personal frustration and submit reports in the format requested which in the long run was... opportunities around you. You just need to be prepared to take on the challenges and be proactive and...

...should be prepared with different questions than those on the prep calls, but often the prep calls provide...published on LinkedIn. Follow Addgene on LinkedIn for repository news and updates. Panel discussions are becoming...nonprofit models that differed from the companies represented on the panel. This "spontaneous" participation...purpose. If possible, make sure the panelists are prepared to stay around to meet and greet audience members...

... a peer-reviewed manuscript OR a preprint. In the case of a preprint submission, we are happy to update...containing instructions on how to prepare your plasmids and a prepaid label with shipping materials to ...journal. To start this process, simply enter your preprint title or a general description of the deposit....set of entry fields to describe the next insert. Repeat as needed for more inserts. When you are finished... propagated in DH5α. For plasmids with highly repetitive sequences (which can be prone to recombination...

...the new plasmids, antibodies, and viral preps in the repository through our Hot Plasmids articles.  Here's...delivered with an enhanced pU6cm promoter, viral replicon amplification of cargo, and heat treatment.  ...//doi.org/10.1038/s41477-024-01786-w. (bioRxiv preprint: https://doi.org/10.1101/2024.02.11.579803).  ...components of many physiological pathways and represent the largest family of human drug targets. To enable...6E10-A5] in Immunohistochemistry (Human). Addgene Report, doi: https://doi.org/10.57733/addgene.6ncijt.  ...

...multiple other labs have reproduced their EGFP findings since the initial report in 2023.)     Figure.... Andrew and Maria were encouraged by previous reports that described magnetic responses of cryptochrome...indicate times when the magnetic field is on. Image reproduced from Frank Hayward et al. 2024 under CC-BY license...), its behavior has already been successfully reproduced by several other labs. Andrew and Maria expect...

...technology that allows precise modifications of DNA (replacements, small insertions, and deletions) without introducing... In our study, we used the TRIP (Thousands of Reporters Integrated in Parallel; Akhtar et al., 2013; Schep...Chromatin Position Effects Assayed by Thousands of Reporters Integrated in Parallel. Cell, 154(4), 914–927...., Raguram, A., & Liu, D. R. (2019). Search-and-replace genome editing without double-strand breaks or ...insertion efficiencies using sequence features and DNA repair determinants. Nature Biotechnology 2023, 1–11. ...context on Cas9-induced DNA double-strand break repair pathway balance. Molecular Cell, 81(10), 2216-2230...

...the sequencing results of the plasmids in our repository with known reference sequences, such as full ... entries in NCBI’s numerous databases. As our repository has grown over the years (we now have over 60,000.... At Addgene, we use blastn to identify any discrepancies in Sanger sequences, such as mismatches, deletions...determine the potential effect of any nucleotide discrepancies. The blastp and blastx programs are optimized...Similar to nucleotide sequences, proteins often have repeated or highly homologous regions, which by default... standard blastx search. An alignment omitting repeated regions can be confusing, such as when you attempt...

..., consider an approach that prioritizes more reproducible/renewable reagents, such as recombinant or hybridoma-derived...cells that are amenable to genetic knockout, this represents a very robust approach to antibody validation...quantified using the Attune NxT Flow Cytometer with representative images showing the staining intensity in the...increase confidence that the labeling observed represents the expression level of the target of interest...antibody clones The antibody validation process represents a high level of effort and time, which may not...evidence is then needed to confirm that the staining represents the target in your sample of interest (bottom... specific detection of human TRPA1. Scientific Reports, 9(1), 18500. https://doi.org/10.1038/s41598-019...

...transformation   Spectinomycin/streptomycin Spectinomycin and streptomycin both inhibit protein synthesis...expensive antibiotic compared to streptomycin. Spectinomycin/streptomycin can be successfully used for plant...offers is marginal. Fortunately, amp can easily be replaced with carb if satellite colonies are an issue. ...spectinomycin). Cons: Cost (spectinomycin) or stability (streptomycin), doesn’t work for all bacteria.   Zeocin  Zeocin...

...transcription) is demarcated by viral genome replication (Harrach, 2014). Given that the early genes ...encapsidate new virions) are not expressed. Note that replication-competent AdV vectors also exist and are used...the final topic, which is that natural AdVs, replication-competent viruses with a recombinant genome, ...(Tollefson et al., 1996) – cell death has been reported when using vectors, and immune inactivation of...pre-existing toolbox for adenoviral vectors in Addgene’s repository is smaller than for other vector types, we hope...Hermiston, T. W. (2003). Multigene expression from a replicating adenovirus using native viral promoters. Mol ...

...purification kits. Kit-free alkaline lysis plasmid miniprep Starting material: 2 mL bacterial culture Product...Product: plasmid DNA This kit-free plasmid miniprep protocol from Addgene follows a similar workflow as ...authors estimate that a gravity-based column plasmid maxiprep kit takes 1-1.5 hours to complete, while this ... minutes. Additionally, while commercial plasmid prep kits come in discreet sizes (mini, midi, maxi, giga....0007750  Vogelstein, B., & Gillespie, D.A. (1979). Preparative and analytical purification of DNA from agarose...Purification and Gel Extraction Kits. Scientific Reports. https://doi.org/10.1038/s41598-018-30316-w  Zou...

...activator such as VP64 (Gilbert et al., 2013), or repression of transcription through binding a promoter region...expression plasmid. Experimental use Since the initial report describing CRISPR-mediated purification of a specific...dCas9, with bound DNA, can then be purified by streptavidin high-affinity purification. This system is both...including Protein A, 1xFLAG, 2xAM, and HA have been reported, as well as affinity purification with an anti-Cas9...chromatin immunoprecipitation (enChIP)." Scientific reports 3 (2013): 3171. PubMed PMID: 24201379. PubMed Central.... PubMed PMID: 28841410. Qi, Lei S., et al. "Repurposing CRISPR as an RNA-guided platform for sequence-specific...