We narrowed to 583 results for: BASE;

...publication: http://www.freebase.com/base/aptamer Cruz-Toledo, J. et al. Aptamer base: a collaborative ...collaborative knowledge base to describe aptamers and SELEX experiments. Database, bas006 (2012). PubMed. For a ...biofactories by demonstrating the use of CRISPR-based RNA scaffolds to mimic natural transcriptional programs... RNA scaffolds follows the rules of Watson-Crick base pairing, allowing for rational, straightforward ...straightforward design. Since the technique relies on base pairing, it has promise to be host-independent, beyond the...the University of Western Ontario studying plant-based recombinant pharmaceuticals. Follow him on Twitter...

...utility of cytidine and adenine base editors that enable C-to-T and A-to-G base substitutions respectively.... were first discovered in bacteria, most CRISPR-based genome engineering has taken place in other organisms...CRMAGE system combines CRISPR and recombineering-based MAGE The Nielsen lab’s CRMAGE system is a fast, ... method that combines CRISPR and recombineering-based MAGE (Multiplex Automated Genome Engineering) technology...Bikard et al., 2013), the Qi lab employed a gRNA-based design more familiar to those using CRISPR in eukaryotes...catalytically dead Cas9 enzyme allows reversible repression based on the presence of the inducer, in this case anhydro-tetracycline...RNAP-Omega-dCas9 fusions. Here’s some others. SoxS based CRISPRa In the search for a CRISPR activation method...

...IDT estimates that there is an NGG PAM every 42 bases in the human genome. SpCas9 targeting should be ...difficult if there’s no PAM near your insertion site. Base editing, another method to create point mutations...Employing xCas9 for transcriptional regulation and base editing Hu et al. also engineered transcriptional...NGG PAM, confirming the flexibility of xCas9 3.7. Base editor variants with xCas9 are similarly flexible...nuclease, xCas9 3.7 fused to third generation cytidine base editor architecture (BE3) slightly improves NGG ...efficiency over that of SpCas9-BE3 (37 vs 28%). Cytidine base editing at NGT, NGC, GAA, and GAT PAMs is improved... with a smaller increase in NGA editing. Adenine base editor xCas9 3.7-ABE also follows the same pattern...

...IBEX Knowledge-Base, with every contributor listed as an author.  The IBEX Knowledge-Base is an invaluable...amplified staining methods, and oligonucleotide-based staining. Of course, if you’re interested in using... Ziv Yaniv, PhD, have created the IBEX Knowledge-Base, a community repository and resource for spatial...open-software community when designing the IBEX Knowledge-Base, seeking to create an iterative process that is .... Everyone who contributes to the IBEX Knowledge-Base is named on the Zenodo release in keeping with their...components of building both the community and the database. They particularly encourage people to upload ...

...plasmids Cytosine base editors with minimized off-target DNA editing The original cytosine base editor, BE3,...research! This time: Optogenetic repression in yeast Base editing CRISPRi in Caulobacter crescentus Multiplexed...Multiplexed gRNA for human pluripotent stem cells CRISPR-based DNA demethylation Optogenetic repressors of gene... Read the paper in Nature Biotechnology Find the base editor plasmids CRISPRi in Caulobacter crescentus...Scientific Reports Find the plasmids Enhanced CRISPR-based DNA demethylation Figure 2: The components...

...that cover some of the hottest CRISPR tools. Base Edit: Base editing plasmids allow you to make precise ...causing double-strand breaks. Our new page shows you base editing plasmids for mammalian systems, bacteria...bacteria, plants, and yeast. Popular cytosine base editor pCMV-BE3 has been requested nearly 400 times, and ... has already earned a blue flame for its adenine base editing capabilities. Epigenetics: CRISPR has made...for Gene Editing Day 9: Precision editing: Single Base Editing with CRISPR and Enhancing CRISPR Targeting...

... on base editing from the first generation of base editors to the eighth generation (and dual base editors...Addgene gave to me: SHERLOCK and DETECTR, two CRISPR-based methods to detect nucleic acids.  On the third day...

... Course-based Undergraduate Research Experience (CURE). Having incorporated a discovery-based lab on planarian...and research-based learning in a year-long biochemistry laboratory : Part II – research-based laboratory...thinking skills (Knutson et al., 2010a,b). Classroom-based research projects have also been proposed as a mechanism...References Bangera, G., Brownell, S. 2014. Course-based undergraduate research experiences can make scientific... E., Kowalko, J., Keene, A. et al. 2022. Course-based undergraduate research experiences (CUREs) as a ...research to students; Using inquiry and research-based learning in a year-long biochemistry laboratory:...Implementation of a collaborative series of classroom-based undergraduate research experiences spanning chemical...

...in the region around the cut site. It shows, base-by-base, the average amount of signal that disagrees...Editing Results with ICE on Synthego’s blog. CRISPR-based genome engineering has revolutionized the gene editing...Sanger sequencing data alongside the analysis of NGS-based amplicon sequencing data and found that the accuracy...enabling a ~100-fold reduction in cost relative to NGS-based amplicon sequencing. To use the tool, you start ...Analyze Complex Edits   Traditional Sanger sequencing-based analysis tools are unable to detect or analyze complex...region around the guide binding site(s). The sequence base calls from the .ab1 file are also shown above each...error-prone repair typically result in mixed sequencing bases downstream of the cut. Summary ICE generates NGS-quality...

...positions to the base expression. When compared to the control — which should be base expression — a peak...with the negative control population (Figure 1). Based on where the GFP- population ends, you can draw ...cells and might be upregulated or downregulated based on your experiment. You can see from Figure 2 that...that determining where to place your gate based on the dot plot would be complicated.  Figure 2...   In such cases, instead of trying to quantify based on percentage positive or negative in your dot plot...populations of interest with normal gates, just based on a histogram plot instead of a dot plot. However...overlaps with the baseline suggesting this sample has a comparable expression to the baseline/negative control...

...of cancer through different mechanisms. New FRET based tools to study epigenetics Epigenetic research uses... Alice Ting’s lab has developed a new, plasmid-based biosensor to visualise changes in histone methylation...

...of fluorescence resonance energy transfer (FRET)-based calcium biosensors. Perhaps most notably, the Twitch.... “It gives you more information than intensity-based readouts.” Griesbeck also points out that the approach...now be applied to dozens of other published FRET-based sensors. For example, his team is now applying it...Universiteit Eindhoven has similar interest in FRET-based sensor proteins, particularly for the intracellular...

...your next experiment. Nucleic acid based machines Recombinase-based State Machines Enable Order-dependent...Roquet from the Lu Lab at MIT developed plasmid based systems that report the order in which specific ...received by a cell. These so-called recombinase-based state machines could potentially be used to report... Read this post to learn more about recombinase-based state machines and how they function. Adapting Toehold... Toehold Switches to Detect Zika Virus The RNA-based toehold switches described in this post are engineered...

... in AAV. A dCasMini-mediated adenine base editor allows for base editing, while gene editing can be done...perturbation by: Gabrielle Clouse While many CRISPR-based genetic screening methods have primarily used Cas9...genetic screening toolkit by optimizing a Cas12a-based method for screens. Cas12a’s ability to perform ...multiple genes in parallel. Lastly, this Cas12a-based screening method was used to create a genome-wide... Rachel Leeson This simple, inexpensive plasmid-based ladder system is a cost-effective alternative to...efficacy to dCas12a systems in both gene activation and base editing applications.  Figure 4: Overview...

...insert genes into plasmid backbones, base pairing between these ~4 base overhangs isn’t strong enough to ...recombination, it also doesn’t require a ligation step. Based on the robust system of homologous recombination...insert are (separately) digested to create 10–12 base “chewed back” overhangs, they can be annealed, forming...preventing T4 from chewing back the entire 10–12 bases. As such, the use of LIC is often limited to specially...overhangs must be accessible to allow for complementary base pairing, so SLIC can’t be used if the overhangs ...compared to Gibson assembly, a cloning method also based on homologous recombination. The higher temperature...

...under non-denaturing conditions separate proteins based on size. Whether higher molecular weight complexes...tendency. Another technique that separates molecules based on size is ultracentrifugation. Sedimentation equilibrium...protein forms dimers. Finally, spectroscopic methods (based on fluorescence polarization) can be used to detect...EGFP. Constantini et al, (2012) developed a cell-based assay that does a better job of detecting homodimerization...whereas it was previously concluded to be monomeric based on gel filtration (Chudakov et al, 2007). We have... and negative controls for proper ER structure. Based on these data, we conclude that mKOk has dimerization...Amsterdam) where she develops genetically encoded FRET based biosensors.     Additional Resources on the Addgene...

...Potential advantages of Cpf1 Type II CRISPR systems based on Cas9 were thought to be the simplest CRISPR systems...fashion, creating a 5 nucleotide 5’ overhang 18-23 bases away from the PAM. Cpf1’s staggered cleavage pattern...potential to eclipse any of the CRISPR nucleases in AAV-based applications. Get More Cpf1 Background Here THE ... Figure 1: Effects of (A) double and (B) single base pair gRNA-target mismatches on modification by AsCpf1...with single and double mismatches throughout the 23-base sequence. Double mismatches ablated Cpf1 activity...were present in the 3’ end of the target sequence (bases 19-23). Cpf1 is also sensitive to single mismatches...et al. saw no decrease in Cpf1 activity with 4-6 base deletions at the 3’ end of the target sequence. ...

...score match. This tool’s algorithm can be adjusted based on query, reference, and shared allele search preferences...called Cellosaurus, a growing cell line information database describing nearly 145,000 cell lines. Cellosaurus...researcher, Cellosaurus was able to both update their database and sent the updated information to the cell line...

...     A new positive selection-based RNAi method Until now, all DNA-based RNAi experiments typically involved...pervasive off-target effects. This conclusion was based upon numerous phenotype discrepancies between gene...considerations present at the first stage of an RNAi-based investigation. Creation of the RNAi trigger requires...considerations, such as the starting material (DNA-based vector or purified RNA) and the initial state of...allows easy creation of long hpRNA with Gateway-based cloning and straightforward positive selection of...occurrence of off-target effects. Nevertheless, amiRNA-based RNAi experiments routinely display highly specific...#137884). Both vectors allow the creation of amiRNAs based upon miRNA precursors of rice and Arabidopsis species...

... pegRNAs. Chromatin-based prime editing efficiency prediction While sequence-based prediction models from...building a chromatin-based prediction tool (ePRIDICT). In summary, machine learning-based prime editing ...way to predict the efficiencies of pegRNA designs based on patterns observed from high-throughput experiments...(used for PE3 and PE5 prime editing strategies) based on a DeepSpCas9 score ranking (Kim et al., 2019)... predicts how well a genomic site can be edited based on its chromatin environment. After selecting the...activity prediction by DeepSpCas9, a deep learning–based model with high generalization performance. Science...

... created a codon adaptation index (CAI) that is based on the codon usage frequency in a reference of highly...improve gene expression by changing synonymous codons based on an organism's codon bias. The idea is that scientists...can make mutations throughout a gene of interest based on an organisms codon usage bias to increase translational...code (Gustafsson et al., 2004), such as Hoogsten base pairs and wobble pairs.  Figure 2: (A) The...different tRNAs, including those that exhibit wobble base-pairing (a tRNA that can recognize multiple synonymous...Chappell SA, Zhou W, Mauro VP (2005) An mRNA-rRNA base-pairing mechanism for translation initiation in ...-0270(02)00362-x Stadler M, Fire A (2011) Wobble base-pairing slows in vivo translation elongation in ...

...a 30 nt guide when C or U base-flips are present across from the target base. However, they saw that in...Cas13 enzymes prefer targets with a given single base protospacer flanking site (PFS) sequence, but orthologs...so they cannot directly edit the genome. A Cas13-based RNA editing system would likely be reversible and...it is sensitive to mismatches in target RNA from bases 12-26 of a 30 nt target sequence. For the editing...targeting work with Cas13, REPAIR is the first CRISPR-based system to enable precise RNA editing. The addition...al. “A cytosine deaminase for programmable single-base RNA editing.” Science 2019 Jul 26;365(6451):382-...RNA Targeting with Cas13a Learn about CRISPR DNA Base Editing Resources on Addgene.org Browse All CRISPR...

...Class 2 CRISPR systems, including the type II Cas9-based system, require a single-component nuclease to mediate...fashion, creating a 5 nucleotide 5’ overhang 18-23 bases away from the PAM. With this information, they turned...advantages of Cpf1 over Cas9 Type II CRISPR systems based on Cas9 were thought to be the simplest CRISPR systems...with single and double mismatches throughout the 23-base sequence. Double mismatches ablated Cpf1 activity...were present in the 3’ end of the target sequence (bases 19-23). Cpf1 is also sensitive to single mismatches...et al. saw no decrease in Cpf1 activity with 4-6 base deletions at the 3’ end of the target sequence. ... Figure 2: Effects of (A) double and (B) single base pair gRNA-target mismatches on modification by AsCpf1...

...previously uncharacterized enhancers. The popular TALEN-based LITE system, which uses light to regulate transcription...deacetylases. CRISPR and epigenetics As with many TALEN-based technologies, the advent of CRISPR has made targeting...strand breaks. Below you'll find some of the CRISPR-based epigenetic modifiers available from Addgene. For...Albert Cheng’s lab developed Casilio-ME, which is based on their Casilio system. This system allows for ...Epigenetic modification is certainly not the only CRISPR-based technology designed to alter gene expression. Fusing...cloning platforms for transgene expression and dCas9-based epigenome editing. Nucleic Acids Research 47:e23...Gersbach CA (2015) Epigenome editing by a CRISPR-Cas9-based acetyltransferase activates genes from promoters...

...of research into COVID-19. While this program is based in Canada, much of the information is applicable...support this pace of research, new, centralized databases and centers to support the sharing of materials...into COVID-19 (like here at Addgene!), and new databases are supporting their access to useful resources...internationally COVID-19 Resources Canada is a database that provides information about reagents, funding...the future of research. To find other COVID-19 databases and resources, check out our growing Twitter thread...

.... Development of this tool required creating a database of common features, and devising rules for identifying...when the match is imperfect. SnapGene's feature database The source of common features was our collection...issue is addressed by adding more variants to the database. Another limitation is that by tolerating mismatches.... To prevent misidentification, we augment the database with new fluorescent protein variants. The future...features? We plan to continue updating the feature database. Powering the plasmid maps on Addgene.org with... which common features need to be added to the database – we will work to fill any common annotation gaps...SnapGene supports “Collections”, which are shareable databases of DNA and protein sequences, the next step is...

...Coselection with Ouabain New CRISPR base editors The popular CRISPR base editing technique uses dCas9/Cas9...specificity of base editing. Kim et al. used natural and engineered Cas9 variants to develop five new base editors...number of available target sites for base editing. For each base editor, they observed editing activity...cytidine deaminase portion of the base editor to create SpCas9 base editors with editing windows as small...off-target effects associated with base editing, Rees et al. created HF-BE3, a base editor containing high fidelity...diseases. The Tol2 Gateway-Compatible Toolbox is based on the original Tol2kit generated by the Chi-Bin...These exciting papers further show the potential of base editing for precise genome modification. Plasmid...

...increasing CRISPR editing efficiency, new CRISPR base editors, DIY DNA ladders. You can find additional...Segement on Ouabain Selection  Hot Plasmid 3: CRISPR base editors with enhanced range and specificity. Listen...Listen to to the Segmennt of Improved CRISPR Base Editors Hot Plasmid 4: pPSU Plasmids for generating DNA...

... the target site. Additionally, the TAM changes based on the species of origin of the TnpB protein, much...access to regions previously unreachable with Cas-based editors. TnpBs have a familiar targeting mechanism... bound, TnpB cleaves the DNA at a position 15–21 base pairs downstream of the TAM, creating staggered ...Even for advanced genome editing approaches like base editing and prime editing (Molla et al., 2021), ...currently unavailable, you can use TEEP — a tool based on animal data. Step 2: Prepare Oligos Oligo... invite researchers to explore and use our TnpB-based genome editing vectors. If you have specific questions...Qi, Y. (2021). Precise plant genome editing using base editors and prime editors. Nature Plants, 7(9), ...

...ex: genome editing), our MUCICAT system which is based on the CRISPR-associated transposase system, INTEGRATE...gene expression is an alternative versus plasmid-based expression which can be unstable and lack precise...application of a multiplex genome editing system based on CRISPR-associated transposition.     References...

... editing (SECURE) base editors Keith Joung’s lab recently found that a cytosine base editor (CBE) with...Covalent tethering of DNA template to Cas9 SECURE base editors Nme2Cas9 CRISPR interference in Candida ...efficiency. Read the paper in Nature Find the SECURE base editors All-in-one AAV delivery of Nme2Cas9 with...

.... “You can think of changing bases to other RNA bases or unnatural bases,” she said. “The possibilities...modular repeats, each of which binds a consecutive base in its RNA target. By changing amino acids, it’s...

...initially recognized as a useful vertebrate model based on their straightforward husbandry and ability to...developmental biology that arose from zebrafish-based investigations. Genetic resources for zebrafish...important insights in developmental biology. Microscopy-based lineage tracing Over a period of 24 hours, the externally...approaches (Keller et al., 2008). This microscopy-based lineage tracing defined a high-resolution map of...from the random mutagenesis screens. Microscopy-based lineage tracing can help illuminate how signaling...during embryo development. Single cell sequencing-based lineage tracing Recently, gene expression in all...rather than novel, abnormal cell types. Sequencing-based lineage tracing can help elucidate the gene networks...

...overview of the general schematic of CRISPR/Cas based COVID-19 testing methods.   Rapid detection of...2019 novel coronavirus SARS-CoV-2 using a CRISPR based DETECTR assay Sensitivity:10-100 copies of RNA per... al., 2020). Detecting coronavirus using CRISPR-based SHERLOCK (Specific ​H​igh ​s​ensitivity ​E​nzymatic... A scalable, easy-to-deploy, protocol for Cas13-based detection of SARS-CoV-2 genetic material (CREST)...on the target regions thus making it an affinity-based method. Due to the mismatch sensitivity of FnCas9...Wadford DA, Chen JS, Chiu CY (2020) CRISPR–Cas12-based detection of SARS-CoV-2. Nature Biotechnology. https...coronavirus SARS-CoV-2 sequence detection method based on CRISPR-Cas12. Cold Spring Harbor Laboratory. ...

...be used for cloning or genome engineering and is based on homologous recombination. It allows for direct...non-selectable sequences, point mutations or other small base pair changes, and the addition of protein tags. ...best for point mutations or changes of only a few base pairs.  dsDNA Substrate dsDNA inserts can be made...the DNA sequence of interest and flank it with 50 base pairs of homology to the targeted insert site (Figure...way is to introduce a C/C mismatch at or within 6 base pairs of the edit site. This is an easy fix, but...the wobble codons, i.e. make changes to the third base pair of the adjacent 4-5 codons that alter the nucleotide... DL. Recombineering: A Homologous Recombination-Based Method of Genetic Engineering. Nature protocols....

... the Cameleon-based FRET sensor D1ER, and GCaMPER. Bonus: There are also Cameleon-based FRET sensors available...kinetic parameters. There are three GCaMP types based on their kinetics: GCaMPf (fast) GCaMPm (medium)...wavelength is insensitive. Additional red sensors based on mRuby (jRCaMP1) and mApple (jRGECO1a) are also...reduce these unwanted variables. Twitch is a FRET-based calcium reporter derived from troponin C. It is ...mechanism-guided design of single fluorescent protein-based biosensors. Nature chemical biology. 2021 May;17...

... of viral particles in a sample and are usually based on the presence of a viral protein, such as p24,...incorporated into a viral particle. Consequently, titers based off of p24 quantification tend to be overestimates...depending on the vector backbone. When using qPCR based methods for viral titration consider using a reverse...by antibiotic-resistance or single cell sorting based on expression of a surface marker, subcloning, expansion...more straightforward approaches such as the FACS-based or colony-forming unit assays, whereas labs that...concentration and titration of pseudotyped HIV-1-based lentiviral vectors."Nature protocols 4.4 (2009):...

...is another very experienced and successful coach based in New York. Another experienced coach and speaker...you. There is also a group The Dissertation Coach based in Illinois. A few of the coaches in this group ...specialty for Michelle Capes at Adeptify Me. While based in Madison, Wisconsin she also works remotely and...recruiter. Chris Humphrey from Jobs on Toast is based in the United Kingdom and scientists there would...content for PhDs and postdocs posted by Toronto-based Jennifer Polk of From PhD to Life. She runs great...interesting and it is in English and Spanish. She is based in Spain. The bottom line – there are lots of people...

...fluorescently labeled probes. Colorimetric and luminescence based detection methods are also commonly used. In all... Since the sequencing gel will separate the DNA based on size, the resulting gel should ultimately contain...correspond to every nucleotide in your DNA sequence, based on which ddNTP was added and when. Compiling the...fluorescently labeled ones allowing for fluorometric-based detection. This increased sequencing speed ultimately...world, one up and coming innovation is nanopore-based DNA sequencing which reads the nucleotides of a ...nucleotides (8). Compared to older technologies nanopore-based DNA sequencing machines are relatively small meaning...error rates (10). It is projected that nanopore-based sequencing technologies could one day sequence an...

...and image for long periods of time.  Are antibody based probes a solution? To address these limitations,...these to tag their proteins of interest. Antibody based probes, such as antigen-binding fragments (Fabs)...frankenbody To combat the issues with these antibody based probes the Stasevich lab used an alternative strategy...strategy to develop a genetically encoded antibody based probe that detects the widely used HA epitope in...CDR grafting to stable frameworks and structure-based framework engineering. Methods, 34(2), 184–199. ...