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... AAV Targeting and manipulating specific, genetically-defined neuronal populations with intersectional...calcium sensors and GABA sensors) Calcium sensors The latest calcium sensors from Janelia (jGCaMP7) continue...potential, GCaMP7s is approximately twice as bright, relative to baseline fluorescence, as the GCaMP7f variant...useful for imaging signaling activity in large populations of densely-labeled neurons because background...all our biosensors inventory, and check out our latest vectors below: pGP-AAV-syn-jGCaMP7f-WPRE (104488...Optogenetics continues to be a useful method for manipulating specific neurons. We recently packaged a soma-targeted...specificity of influence measurements by reducing photostimulation of the axons and dendrites of non-targeted...

... can rupture E. coli spheroplasts to release encapsulated proteins. Video by ACS. For a more cartoony ...thousands of times by propagating through an enormous lattice of identical subunits. A brief history of R bodies...coli in the 1980s  (Kanabrocki et al 1986). Much later, it was shown that putting the operon under a strong...mutants that couldn’t extend at a normal pH and later realized that these mutations clustered in a particular...their ability to penetrate other barriers (like vasculature or extracellular matrices) in response to pH ... response to environmental conditions. More speculatively, R bodies might endow divergent organisms with...Protein Piston That Breaks Membranes to Release Encapsulated Cargo. ACS Synth. Biol. (2016). doi:10.1021/...

...proteins regulate vital cell and tissue processes, but their low abundance, complex regulation, and context-dependent...can include background autofluorescence from the plates or media, which can obscure weak signals; photobleaching...expression and localization. HiBiT-tagged targets in translational research Researchers can incorporate HiBiT with...difficult-to-study genes. Its main function is to regulate DNA repair, senescence, and apoptosis. Work by...at R273H to match a mutation frequently found in late-onset GC. HiBiT-TP53 expression was detected by ... of cystic fibrosis transmembrane conductance regulators (CFTR) (Kamada et al., 2023). CFTR is a membrane...., Robers, M. B., Benink, H. A., Eggers, C. T., Slater, M. R., Meisenheimer, P. L., Klaubert, D. H., Fan...

...is software product management, and how does it relate to science? In the simplest of terms, a software...more about the stakeholders, the product managers collate and organize that information and form it into ...science backgrounds, most did not. It was my job to translate complex lab operations workflows into a format... help improve their work. Then, I distill and translate that valuable information back to the rest of ...science companies, try to find internships with translatable skills: working with data, working with people...jargon and know the concepts as soon as possible Later in your career:  Show demonstrated skill of people...project management  Bolster your skills in strong relationship-building and influence. You’ll need a large ...

...gene a megabase away, or looking at the histone methylation status at a double-strand break that needs repairing...chromosomes interact with each other. Chromatin regulation is complex and influences--directly or indirectly...difficult to consolidate every conceivable chromatin-related plasmid in a collection that grows on a daily basis... Yi Zhang Leonard Zon  You!     Chromatin-related genes with plasmids available from Addgene The ...directly on our website. In-exhaustive Chromatin-related genes with plasmids on Addgene.org AGO1 BRCA1...from Addgene Many articles reporting chromatin-related research describe the production of plasmids that...2018 Feb 13. pii: 1722491115. Addgene blog posts related to chromatin This is not the first Addgene blog...

... in an unregulated fashion and thus a marker for transfection. The other (mKate2) is regulated in the ...Constructs using the photodimerizer system CRY2-CIB1 to regulate botulinum neurotoxin were also created. The depositors...new light on mechanisms of miRNA-mediated gene regulation Article contributed by Michael Lemieux  Listen...microRNAs (miRNAs) are recognized as important regulators of post-transcriptional gene expression across...several years of research, the mechanisms of miRNA regulation have yet to be fully elucidated. Ron Weiss’s ... present, it will repress expression of mKate2 relative to EBFP2. Weiss’s group used these reporters to...define the different cells present in a mixed population, or identify differentiation and disease states...

...no better place for a scientist to make new relationships than at scientific conferences. Conferences ...this is a great ice breaker. Of course, real relationships go further and grow over time, but being interested...fun, memorable meal is a great way to build relationships. Make a list of experimental problems or questions...work – casual socializing is how people build relationships that are memorable. It isn’t always appropriate...overdo it (some of my best studies were designed over late night poker games at Gordon Conferences). You don... of connection. You can also see and offer congratulations on job changes and promotions.  Follow academic...scientists on ResearchGate and/or via Pubmed alerts. Congratulate your connections when they get a new paper published...

...ddifferent cell types, including difficult-to-manipulate primary cells, achieving knockdown within 1-2...methods for TRIM-Away delivery. For small cell populations, co-microinjection of TRIM21 protein and the ...the target antibody works well. In larger cell populations, you can deliver these components together using... electroporated cells re-adhered to the culture plate within 4 hr of treatment. In some primary cell lines... CRISPR/Cas9-based system to study epigenetic modulation. While catalytically inactive cas9 (e.g. dCas9...-minute analysis of the kinetics of chromatin regulation and subsequent mathematical modeling. Its reversibility...amount of noise in reporter gene expression by modulating the induction of transcription factors 1 and ...

... in neurons reveals a mechanism for regulating its translatability." Science 343.6169 (2014): 419-422....Therefore, I set out to study how protein turnover is regulated in neurons. Protein turnover is the combined ...the abundance of proteins in cells. Homeostatic regulation of protein abundance is essential for normal ...acids. The use of radioactive materials is highly regulated, requires designated work areas and come with ...proper microscope setup we can observe the whole population of one particular protein in neurons or in any... has recently published techniques to follow translation in living single cells. However, before CRISPR-Cas9...synthesized proteins of interest were detected and later visualized by FUNCAT-PLA. Before these methods ...

...or 16S rRNA isolated from the microbial community) reveals the RBS and allows translation of the reporter... Thus, cells can be both sorted using FACS and isolated with antibiotic selection. The team tested this...followed by a reporter gene. The hairpin blocks translation by sequestering the RBS, but binding of the “...relevant host biomarkers. The lab validated their platform on clinical stool samples by comparison to qPCR...Betaproteobacteria, and Gammaproteobacteria species isolated from different bee species. BTK can be used to...division in the gut microbiota Monitoring microbial population dynamics in the gut can provide insight into ... al. "A low-cost paper-based synthetic biology platform for analyzing gut microbiota and host biomarkers...

..., and electroporated them with an ssODN repair template in induced pluripotent stem cells made from the...fertilization. Since Ma et al. designed a repair template with two synonymous mutations to distinguish it...were able to distinguish between repair from the template vs repair from the maternal MYBPC3 allele. Interestingly...from the single-stranded oligonucleotide (ssODN) template supplied by Ma et al., but instead from the mother...supplied single-stranded oligonucleotide (ssODN) template. Embryos also underwent editing through the NHEJ...NHEJ pathway. Created with BioRender.com. In later characterization studies, the edited embryos proceeded...embryos to undergo editing using the provided ssODN template. In fact, this finding led the authors to title...

..., a researcher can cause them to interact by stimulation with blue light.  Similarly, light can also be... and is reversible upon termination of light stimulation. CRY2clust enables faster assembly and disassembly... and better distribution of clusters without accumulation in subcellular compartments such as nuclear ...Furthermore, CRY2clust, when integrated into a Ca2+ -modulating optogenetic system (OptoSTIM1), caused two-fold...Listen to the soCoChR podcast segment Optogenetic stimulation is a powerful tool used by neuroscientists to...artifactual PSCs commonly observed with direct stimulation are largely absent in the soCoChR-expressing ...rescued by the xanthine produced via complete demethylation of caffeine. For example, the Barrick lab was...

...and Sibernetic, a fluid mechanics simulator that can be used to simulate muscle tissue as it applies to ...of mRNA are important for post-transcriptional regulation in C. elegans. To study the role of 3’UTRs, the... C. elegans 3’UTRs structures, alternative polyadenylation for all protein coding mRNAs, and annotations...GAL-4 based bipartite expression system that can regulate gene expression in C.elegans. In this system a... to activation sequences upstream of genes to stimulate expression (Wang et al., 2016). In 2018, the lab...vectors that each contains a guide RNA and repair template of your choosing (Schwartz and Jorgensen, 2016...PMC3531189. Mangone, Marco, et al. "UTRome. org: a platform for 3′ UTR biology in C. elegans." Nucleic acids...

...Because of the availability of tools for genetic manipulations, mice have become the model of choice for many...experimenter blinding to avoid bias Genetic manipulation in behavioral neuroscience using viral vectors... is often done in conjunction with a genetic manipulation so that behavior can be linked to a specific... region. When it comes to making the genetic manipulation, there are two main decisions to make: 1. Which...inhibition upon light (or drug) application.  Genetic manipulation can also reduce expression levels of certain...spatial and temporal control of gene expression manipulation. They are injected using a stereotactic frame...depending on whether it is an overexpression gene, downregulation via RNAi or gene editing/deletion using CRISPR...

...them in vivo and in vitro, especially in the mid-late 20th century with the advent of hybridoma technology...recombinantly append their proteins of interest with unrelated peptides that already had high quality antibodies...an amide group added to its C-terminus post-translationally, which turned out to be critical for antibody..., another group of researchers was working on isolating monoclonal antibodies to better probe the functions... c-Myc protein or the C-terminus, Evan, et al. isolated six different monoclonal antibodies (Evan, et ...sequenced the variable domains of Myc1-9E10 in the late 1990s (Fuchs, et al. 1997; Schiweck, et al. 1997...Evan GI, Lewis GK, Ramsay G, Bishop JM (1985) Isolation of monoclonal antibodies specific for human c-...

...popular tools because: 1) their genomes are easy to manipulate, 2) they have long-term expression; and 3) they...research is the lack of a method for genetically manipulating neurons throughout the whole brain. Neurons ...Cre-expressing tissues. This inversion creates a template that’s suitable for PCR amplification using pre-designed...Cre in a cell type of interest, a scientist can isolate DNA from the infected animal and use the pre-designed...that cross the barrier can be identified by PCR later. The PNS presents a different problem: it’s spread...measured via GFP expression, was determined 3 weeks later. Table was derived from figure 2e in Chan et al....measured via GFP expression, was determined 3 weeks later. Table was derived from figure 3b in Chan et al....

...purified protein not always available Calculate relative quantification Normalizing to ubiquitously...variation. Confirmation of sequence can be done from relatively cheap plasmid sequencing. While rAbs are monoclonal...to run a western blot. It allows you to detect relatively low levels of antigen, while providing flexibility...assay design. Conjugated secondary antibodies are relatively cheap and easy to source, making it easy to switch...lower concentration of blocker, you can also use gelatin.  Protein-based blockers are inexpensive and easily...it to create a standard curve on your blot to calculate the amount in your sample. (Do ensure your blot... not available in a pure form, you can perform relative quantification, which measures protein amounts...

...have evolved over the years to keep up with the latest tools. In Addgene’s early days, we would ‘spot-...Falls sized rainbow   Over the years we have accumulated more than 300,000 individual nucleotide sequences...help us work in large formats, such as 96-well microplates, and thanks to our dedicated product and development...pipeline. After reviewing and testing several platforms for engineering pipelines, we selected Nextflow...appropriate place in our workflow without having to translate or convert programs to a uniform language.  The...running on a computer, so it safely operates in isolation, and can be easily shared with others without ...concepts and vocabulary of Nextflow so that we could translate our pipeline steps into processes and then connect...

... other molecular techniques to introduce time modulation of the TEV cleavage and improve utility of the...systems at Addgene! Schematic of the evolution platform to generate TEV protease variants. Catalysis by... transcription factor to enter the nucleus and modulate transcription of the mCitrine gene. mCherry is...pMAL-T-Avi-His/BirA, a vector commonly used for in vivo biotinylation via BirA and AviTAG. pMAK contains the signal...competent E.coli (NEB) using IPTG induction. To biotinylate the nanobodies, the authors used biotin dissolved...higher concentration, and results in uniform NB biotinylation. Once induced, the NBs could be easily purified...and gene activation (CRISPRa) are great ways to modulate the expression of nearly any gene via specific...

...inferred sequences present in your edited population and their relative representation in the edited pool. The...editing - enabling a ~100-fold reduction in cost relative to NGS-based amplicon sequencing. To use the tool...guide RNA sequence you used. The ICE tool will calculate overall editing efficiency and determine the profiles...different types of edits that are present and their relative abundances.   The ICE tool also provides a Knockout... DNA from both edited and unedited (control) populations is PCR-amplified and Sanger sequenced. ICE compares...during generation of the ICE Score, the Pearson correlation coefficient (r) is also computed and reported...distribution of indel sizes in the entire edited population of genomes. Hovering over each bar of the indel...