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...the end if you ever need to double-check your vocabulary! A brief review of base editing tools First, ...an inhibitor of base excision repair (BER), the cellular process that catches mutated U’s in DNA and ideally...Researchers in the Xueli Zhang lab were making similar observations around the same time. (The papers ... journal on the same day.) This team created a similar construct that they called a Glycosylase Base Editor...the base editor. Black arrows indicate natural cellular processes. Created with BioRender.com. Beyond...abasic site. But how that site is filled depends on cellular factors. Multiple labs tried to identify beneficial...repairing a GBE-derived abasic site requires more cellular factors binding to DNA. However, ABEs were improved...

...antibodies may have similar emission spectra, meaning they will emit fluorescent light at similar wavelengths ...fluorescence for both at similar levels, meaning that your populations will ‘look’ similar on your flow plot....between the two of them because they emit at a similar wavelength. However, if your flow cytometer reads...is if the names of the fluorescent markers are similar, you will need to compensate between them (although...against each other, you should be able to draw perpendicular lines that intersect your populations. If you...population needs to be shifted, consider your perpendicular lines. In this case, the population on the left...needs to shift towards the X axis to become a perpendicular vertical line.     In general, most spillover...

...associated with cellular organelles, scientists have been able to study many cellular processes, including... for studying the organization and function of cellular systems. FPs have been used to track protein localization... localization, cell structure, intracellular trafficking, and protein turnover rates. Additionally, by...trafficking. Although FPs have enabled discovery of many cellular mechanisms, there are some limitations to working... ultimately misinterpretation of the protein’s cellular role. One way to avoid the pitfalls of overexpressed...SapTrap The Jorgensen Lab recently developed a modular plasmid assembly toolkit called SapTrap to improve... with C. elegans, several groups have designed modular toolkits to assist with tagging genomic loci in...

... propensity of a particular type of molecule to form a dimer depends on its molecular affinity, termed...likely molecules of a particular type will interact. A protein with a nanomolar Kd will exist primarily...in cells while a protein with a high micromolar or low millimolar Kd is unlikely to form dimers in a cell... fused to a dimerizing FP has potential to form large stable polymers of dimers. 3) FRET biosensors that...dimerizing FP is fused to the POI. These structures are large (microns), bright, non-physiological, and difficult...the papers describing the FPs above including molecular sizing columns and native gels fail to detect ...vitro. Assays to detect FP dimerization in in vivo cellular contexts, such as the CytERM whorl formation assay...

...better access to intracellular targets. If your target is already on the extracellular side of the membrane...structure of your samples, the relationships between cellular components, and the accessibility of your target...cross-linkers work by (as the name suggests) cross-linking cellular proteins. Cross-linking is a chemical reaction...great for preserving your samples and maintaining cellular and tissue morphology. Formaldehyde is the most...ethanol, work by dehydrating and precipitating cellular components. This method of fixation is good for...Permeabilization What is the point?  Antibodies are large proteins that need a little help crossing cell membranes...is suitable for antibodies that bind to the extracellular portion of a protein. B) Permeabilizing with...

...) antibody-dependent cellular cytotoxicity (ADCC) c) antibody-dependent cellular phagocytosis (ADCP). ...die.  Antibody dependent cellular cytotoxicity (ADCC) Antibody dependent cellular cytotoxicity (ADCC) is... If the pathogen is a large parasite, ADCC can be induced through a very similar interaction between IgE...cytotoxins via degranulation.  Antibody-dependent cellular phagocytosis (ADCP) So far, our antibodies have...most famous for is eating. In antibody-dependent cellular phagocytosis (ADCP), Fcγ receptors on macrophages...modulators, shaping the immune response by driving particular cells or responses. And there are still antibody....2007.11.004 Xu Cao, et al. Promoting antibody-dependent cellular phagocytosis for effective macrophage-based cancer...

...and is particularly excited to hear about novel developments in the fields of gene and cellular therapies...(e.g. a postgraduate degree in law, an MBA, or similar), there are many roles out there that require exactly...to manipulate cells (manufactured by one of the larger biotech companies), and as far as I remember, it...experience. With this valuable training behind you (and largely only that is necessary), there are opportunities...Technical Support Advisor, Technical Correspondent or similar. While there isn’t a clear distinction between ...candidate requirements (and maybe also the proposed salary range) should help. In smaller companies, technical...primarily office based, and email/phone are the most popular ways to communicate with customers. Online live...

... several of our most popular CRISPR pooled libraries and one of our most popular barcoding libraries as...formation titering assay or similar protocol. After functional titer is determined, larger lentiviral preps can...depositing lab. (For our more popular libraries, you might receive a similarly-sized aliquot from an amplification...first step will be to amplify the library. This is similar to amplifying a plasmid: transform E. coli with...place to start is with a protocol for a library of similar size and backbone.  Verification Once your library...representation of your amplified library should be very similar to that of the original library. Like amplification...plasmids in the library.  Spinfection If you have a large number of cells to transduce, you may want to consider...

...of neuronal sub-cellular structures GFP is limited in its ability to label subcellular structures in the...reliable primary antibodies for their detection, particularly when antibodies to the POI are non-specific,...discussed above, weakly expressed proteins in particular are difficult to track with these classic probes...researchers in Loren Looger’s lab have developed new molecular tags combining the advantages of both (Viswanathan...used to amplify the direct smFP signal within sub-cellular structures. For instance, smFPs have been used... the stratum oriens and the stratum lacunosum moleculare were better resolved with smFP_FLAG than GFP....ability to distinguish weakly expressed and/or similar proteins Through their multiple, highly specific...

...use different RT enzymes and performed large edits with similar or better efficiency than PEmax, the current...Optogenetic proximity labeling and TransitID Tracking cellular history with protein ticker tape Teaching an old...REcipient Cells Through Envelope Design). This modular approach combines an interchangeable set of fusogenic...and in vitro applications.   Figure 2: Modular features of DIRECTED particles where the fusion... et al. (2023). Cell type-specific delivery by modular envelope design. Nature Communications, 14(1), ... TurboID and APEX2, are localized to different cellular compartments. They showed APEX2 can use an orthogonal...sequential labeling by TurboID and APEX2 at different cellular locations. C) Enrichment of dual-labeled proteins...

...Covalently to DNA via a Phosphotyrosyl Linkage.” Molecular and Cellular Biology 5(11): 3274–79. PubMed PMID: 2427927...blogger Katrin Michel. Cre-lox is an incredibly popular and powerful site specific recombinase (SSR) system... and Sadowski 1985). The FLP system works in a similar fashion to the Cre system. The FLP recombinase ...excision, inversion, and translocation of DNA in a similar fashion as between loxP sites (For details see ...mammalian cells (usually grown at 37 °C). Smart molecular evolution led to the identification of FLPe which...by adding a nuclear localization sequence (SV40 Large T nuclear localization sequence) and expressing ...codon optimization resulted in FLPO, which shows similar efficiency to Cre (Kranz et al. 2010). These differences...

...fluorophores that have a similar brightness - even though in the most popular pair, CFP has five-fold ...technique has revolutionised the way we apprehend molecular complexes and is still a very useful tool.    ...living cells using a microscope. Historically, molecular interactions were detected by indirect means often...FRET reduced the scale of our perception about molecular interactions. What is FRET? In their JCB 2003...is transferred non-radiatively from an excited molecular fluorophore (the donor) to another fluorophore...fluorophore (the acceptor) by means of intermolecular long-range dipole-dipole coupling." The emission of the acceptor... Constructs to target mTurquoise2 to various subcellular compartments pCEP4CyPet-MAMM Cyan Mammalian...

...Eindhoven has similar interest in FRET-based sensor proteins, particularly for the intracellular imaging of...effort to understand the connection between molecular- and cellular-level events and behavior. It’s a problem... want them to go. Griesbeck is motivated by a particular interest in monitoring the activity and biochemistry..., Griesbeck adds that this color change has a particular advantage for observing events taking place inside...optimized by testing various linker configurations in a large-scale functional screen in bacteria. Sensor variants...quite a robust way to make a FRET sensor with a large dynamic range,” he says. Merkx recently applied ...Hot Plasmid page, where we highlight some of the popular & newly available plasmids at Addgene. If you're...

...salts. Capillary electrophoresis Quantifying DNA using capillary electrophoresis is similar to quantifying...first before larger fragments. Figure 1: The quantification of a RNA ladder using capillary electrophoresis...Visit the Molecular Biology Reference Find plasmids for your research Browse protocols for molecular biology...electrophoresis. Smaller fragments migrate quicker than larger fragments. These fragments are quantified over ...time. Smaller fragments migrate more quickly than larger fragments. Image from OpenWetWare. This method...resources on the Addgene blog Read more about molecular biology protocols and tips Find the best way to...

...unuseable. With the large size of the SpCas9 nuclease (~4.2 kb) this is a particularly prominent problem... Numbers in the large colored circles are rough approximations of the total number of CRISPR plasmids...plasmids for that particular organism available at Addgene. Percentages represent the fraction of that total...function. One huge reason CRISPR has become such a popular genome editing tool is its developers’ willingness...that, for many groups, may not be worthwhile. Particularly if in the end all they want to do is to quickly...of insert) and the reduced titer associated with large inserts can make lentivirus production frustrating...libraries now available in lentiviral format Another popular set of tools in the repository are CRISPR pooled...

...and more. MoClo modular cloning system Synthetic biologists have developed a modular cloning strategy... Victor de Lorenzo's lab has engineered a modular mini-Tn5 vector that can be used to generate random...mechanism for sgRNA delivery and providing a means for large scale functional screens. For more information on...the cloning process for complex DNA constructs, particular for those involving recombination elements such...such as FRT or Lox sites, and provides a modular system for easier exchange and re-use of existing elements...provides improvements to classical cloning methods, particularly for complex transgenesis constructs. Kirchmaier... kinases Receptor tyrosine kinases (RTKs) are a large class of cell-surface receptors which play a critical...

...binding partner avidin are commonly used today in molecular biology for an array of different techniques and... interaction and the uses of biotinylation in molecular biology! Learn about in vivo biotinylation of ...biotin is most commonly known for its role in molecular biology, biotin (Vitamin H or Vitamin B7) is first...carboxyl groups to organic acids to produce important cellular metabolites. Biotin acts as a carboxyl carrier...temperature or organic solvents. Uses of biotin in molecular biology Biotin is a relatively small, water soluble...biotin and its strong interaction with avidin for molecular biology and biotechnology purposes.  Once a polypeptide... or co-expressed with avidin or a structurally similar protein that was discovered in the 1960s, strepavidin...

...of polylinker sequences of molecular vectors". Proceedings of the Belarusian State University Series of...for allelic exchange Read our molecular biology reference Watch molecular biology protocol videos ...contain sacB and will not grow on sucrose. One popular counterselection vector is the pDS132 sacB-based...counter-selection. Anecdotally, there is also a large amount of user-to-user variability, and in some ...colonies turn subtly blue or magenta. This is particularly valuable when first optimizing conjugation in...modification of bacterial genomes. It can accomplish large deletions, insertions, and even precise point mutations...Harvard. Jake received his MD and a PhD in Cell and Molecular Biology from the University of Pennsylvania, and...

...federal regulators. But the speed is impressive on a larger scale, too. The first paper reporting genome editing...later, an A→G base editor (ABE) followed based on similar principles (Gaudelli et al., 2017). These editors...would be tested for their base-editing efficacy in cellular models of KJ’s CPS1 mutation. Every genetic target...base editors that could target KJ’s mutation in cellular models tested both the SpG and SpRY variants of... of Cas9. With many PAMs available, a large number of gRNAs could be screened for the most efficient editing...PAM preferences (Silverstein et al., 2025). In cellular models of KJ’s mutation, SpG Cas9 + ABE8e performed...Up to 5 bystander adenine bases were edited in cellular tests, in addition to the target base. However...

...beta subunits. Both subunits have short intracellular ends and large ectodomains containing ligand-binding...signaling pathways and the linkage of intra- and extracellular environments. Found on nearly every cell in ...unique antibodies that bind discrete subunits or particular subunit pairs. Some of the antibodies in the ...the cell membrane, physically connecting the extracellular environment and the cytoskeleton. Most integrins...affinity for ligands. When integrins interact with extracellular ligands, their ectodomains can extend and open...mediating cell adhesion and migration, initiating intracellular signal cascades, and allowing agile responses...it is resisted by the ligand embedded in the extracellular environment. This further stabilizes the extended...