...dCas9-based tool for DNA demethylation. AAV Q&A with Tim Miles: While not technically a research story, I ...couldn't resist adding in this excellent Q&A from Tim Miles on everything from AAV prep to targeting to...

...AAVs” with the director of CalTech’s CLOVER lab, Tim Miles, PhD, we invite you to head on over to our ...our YouTube channel and watch it now! (Bonus: Tim kindly offered to finishing answering all submitted questions...

...students. Many thanks to our guest blogger Tim Herman! TIm Herman is the director of the MSOE center for...This post was contributed by Tim Herman, director of the MSOE center for BioMolecular Modeling and the...become real”.  We sometimes forget that the models we create are around 30 million times larger than the...Genome Visualization Tips for a First Time CRISPR User by a First Time CRISPR User Resources on Addgene....

...former colleagues! I ran into past ASM President Tim Donohue (see below) on my way back from getting a...Research Center and a fellow Rhodobacter enthusiast, Tim also hosted me at his University of Wisconsin lab... annual meeting (ASM Microbe 2016) for the first time. Our goals at the meeting were to network with scientists...this Broad Institute lab is doing creating well-optimized CRISPR screening technology. Addgene got several...

...CapsidsResearchers: Cynthia Arokiaraj; Viviana Gradinaru; Tim ShayMultifunctional Neuronal Fibers: Modulation from...grateful for your writing, your knowledge, and your time. ...

...Gene Type Wildtype Mutant Partial Sequence Codon-Optimized gRNA Other Tags GFP FP other Flag GST His TEV ...Gearhart 44333 pMX_mTBP TBP LTR Parkinson's Marc Timmers 44978 GST-FUS-WT FUS GST tac ALS Robin Reed 44981...

...enriched lentiviral CRISPR libraries developed by MIT's Tim Wang and colleagues, including David Sabatini and...CRISPRs was limited to modification of a few loci at a time,” Wang said. “It was very difficult to screen in...

...without the artifacts sometimes associated with plasmid-based overexpression. Tim Formosa, at the University...—in frame, of course—and start tagging! Yeast-optimized fluorophores for imaging Lee S, et al. (...series of blue, green, and red FPs that are codon optimized specifically for expression in yeast. These tagging...Based on their findings, the authors recommend optimal FP combinations for use in yeast imaging, categorized... output requirements. Select from these yeast-optimized fluorophore tagging vectors for your single or...

...cyto).iATPSnFR2.S29W.A95K.HaloTag AAV5 Biosensor Tim Brown New viral prep pGP-AAV-syn-flex-iGABASnFR2...

...There are digital and analog versions of each, and Tim Lu even compared his "memory" gene codes to a Turing...

...conjugate or conjugation process. Sodium azide, an antimicrobial agent, can block some amines needed for conjugation...In those cases, thimerosal can be used as an antimicrobial agent. Other chemical interferences with conjugating...measurable light or color in response to a specific stimulus, can be conjugated to antibodies in order to allow...produces a colored product that accumulates over time which is relative to the amount of enzyme present...purchased already conjugated, which can save significant time in the lab. However, if the antibody you want is...ubiquitously expressed protein such as GAPDH or tubulin (sometimes referred to as housekeeping genes). This allows...protein or protein interactions in a specific timeframe or context. Immunoprecipitation (IP) IP uses immobilized...

...neurons upon exposure to light through direct stimulation of ion channels. Naturally occurring channelrhodopsins... nonspecific cation influx into the cell when stimulated. Channelrhodopsin-2 (ChR2), the first widely ...been created via genetic point mutations, codon optimization, and chimeric fusion of domains from two different...engineered with enhancements such as: Human codon optimization Example: Halo Increased photocurrent amplitude...activate different neural populations at distinct times. Temporal considerations. Temporal precision is ...and promoter system used, there is an incubation time (days to weeks) until peak expression of the opsin...fused to Cry2 and the other half to CIB1, light-stimulated heterodimerization can reconstitute the protein...

... an optimized gRNA for your application (see: Plan Your Experiment ). In addition to an optimized gRNA... currently no defined measure of fidelity, so sometimes they will be referred to as increased fidelity...structured motif to enhance pegRNA stability PEmax - optimized PE architecture; increased synergy between PE ...compared to HDR. Prime editing requires more optimization and typically has lower efficiency (< 50%) than...integrase landing site ( attB ) and the other to stimulate DNA repair. This can result in more efficient .... gRNA design for CRISPR libraries is usually optimized to select for gRNAs with high on-target activity...activators and targeting promoter regions. You might sometimes see these referred to together as CRISPRi/a. dCas9...

...field of recombinant DNA technology have been optimized for their use of studying and manipulating genes...bacterial cell division is reduced (i.e., it takes more time to copy this extra DNA). Because of this reduced...form of another antibiotic resistance gene (this time, under the control of a non-bacterial promoter) ...origin of replication, and MCS. They are small and optimized to help in the initial cloning of a DNA fragment...developed the process termed Sanger sequencing, sometimes referred to as chain-termination sequencing or...Sanger sequencing requires a lot of DNA because the ultimate goal is to have a fluorescently tagged nucleotide...captures the fluorescent color that is emitted each time a base is added. Again, each base (A,C,T, or G) ...

... over the years to move around pieces of DNA. Oftentimes several approaches will work for any specific...specific DNA sequence, which can make it difficult at times to identify compatible restriction enzymes, that...behind a short scar in the DNA sequence and can be time consuming compared to other cloning methods. You...simplifying the process and reducing the amount of time required for cloning. To utilize this approach, ...proteins. Thus, you can clone your gene of interest one time into a donor plasmid (or acquire one that already...polymerase used, and the single A overhangs degrade over time, further reducing ligation efficiency. TOPO® ready...

...putative target sites have been identified, it is time to choose which site is likely to result in the ...Using validated gRNAs can save your lab valuable time and resources when carrying out CRISPR experiments...Once you have selected your target sequences it is time to design your gRNA oligos and clone these oligos...proceeding with your experiment, it may be necessary to optimize your delivery conditions. Learn more about CRISPR... gRNA and Cas enzyme to your target cells, it is time to validate your genome edit. CRISPR editing produces...

...vector into which one ultimately wants to ligate the insert. Often times it is less time consuming and easier...experiment, the important component to consider and optimize is the transfer plasmid. 2nd generation lentiviral...post‐transcriptional regulatory element; sequence that stimulates the expression of transgenes via increased nuclear...

...set of anti-HA frankenbodies (Zhao et al. 2019), Tim Stasevich and Hiroshi Kimura’s labs have recently...Frankenbodies for modular imaging of cellular targets Optimized genetic screening technique using Cas12a for multiplexed...https://doi.org/10.1038/s41467-019-10846-1.   Optimized genetic screening technique using Cas12a for multiplexed...have expanded the genetic screening toolkit by optimizing a Cas12a-based method for screens. Cas12a’s ability...increase the potential of Cas12a genetic screens by optimizing enzyme activity using an enhanced Cas12a variant...from DeWeirdt, et al. 2021 To validate this optimized genetic screening technique, researchers identified...simplicity for combinatorial genetic screens, and this optimized toolkit furthers Cas12a’s potential in future ...

... to a startup Mike Koeris and fellow co-founders Tim Lu and others first came up with the idea that would...raised $6 million. The biggest challenge during this time was learning to communicate effectively to non-scientific...complications and potential conflicts that waste money and time. Most importantly, “the company does not become ...become real until someone takes the risk full-time and abandons all safety nets.” Raising seed money. “Before...formerly VP Operations of Sample6) for taking the time to speak with us about the biotech startup process...

... indicating the optimal serotype(s) for transduction of a given organ. Tissue Optimal Serotype CNS AAV1...al. , 2015 from the Zhang Lab to find plasmids optimized for use in AAV, or containing Staphylococcus aureus...

...Currently, Aggarwal, and colleagues Filip Tomaska and Tim Hanson, are characterizing variants of iGluSnFR3 ...Campus, HHMI where he works on developing and optimizing biosensors in the lab. He has recently developed...sensory input from our extremities. “Here, painful stimuli are processed by a network of excitatory and inhibitory... demonstrate pain response due to blue light stimulation of dual virus-expressing CR neurons. (C) Duration...biosensors.   This screening method allowed Tian to optimize a red‑shifted archaerhodopsin‑derived genetically...

...24336571. PubMed Central PMCID: PMC4089965. 9. Wang, Tim, et al. "Genetic screens in human cells using the...References 1. Cheng, Allen A., Huiming Ding, and Timothy K. Lu. "Enhanced killing of antibiotic-resistant...PMCID: PMC4151723. 2. Doench, John G., et al. "Optimized sgRNA design to maximize activity and minimize...

...Plants. Carola Engler, Mark Youles, Ramona Gruetzner, Tim-Martin Ehnert, Stefan Werner, Jonathan D. G. Jones...to overcome them, and why she spends some of her time engaging with others on Twitter, among other things... transcripts as the ones that people use all the time. Now there are many, so if you want to make a 10...switching to Golden Gate is a conceptual barrier. Sometimes people don’t know how to start. They might want...Most Sainsbury scientists are active on Twitter. Sometimes it’s a good way to find out what’s happening in...experiences, articles about women in science, and the sometimes stupid or uncomfortable things that have happened...say to our readers? Patron: Remember plants. Every time you look at your plate, think about how much that...

...10.1093/nar/gku936 Eva K. Brinkman, Arne N. Kousholt, Tim Harmsen, Christ Leemans, Tao Chen, Jos Jonkers, Bas...into your target cells with Cas9. Hooray! Now it’s time to make sure your genome edits went according to...within a given genomic window, along with the estimated editing frequency of your sgRNA-Cas9 combo. Pro... from the donor DNA. This method can precisely estimate editing frequency in a bulk population and also...Cas9 or gRNA is the issue, check out or blog on optimizing this step, or consider enriching for transfected...

...Boston Children’s Hospital and Harvard-based lab of Tim Springer, the founder of IPI. Caitlin Faulds is a...

...post‐transcriptional regulatory element; sequence that stimulates the expression of transgenes via increased nuclear...

...RNA polymerase transcribes DNA to mRNA which is ultimately translated into a functional protein. Thus the...

...G-protein and activate neuronal firing through stimulating phospholipase C, which releases intracellular...

...27760321. PubMed Central PMCID: PMC5081405. 6. Wang, Tim, et al. "Gene essentiality profiling reveals gene...Karolinska Institutet, in Sweden. It can be very time consuming to design 5 guide RNAs (gRNAs) targeting... failing to upregulate the protein following stimulation would then be separated by sorting, and subsequently...much, showing that we are far from consensus on optimal gRNA design. With Green Listed you can also easily...PubMed PMID: 28414855. 2. Doench, John G., et al. "Optimized sgRNA design to maximize activity and minimize...

.... Other molecular targets under research include TIM-3, LAG-3, STAT-3, EGFR, CD20 and CD73 on specific...chemical signals such as IL-8, IFNγ, and colony stimulating factor 1 (CSF-1). Additionally, tumor cells recruit...cell-surface proteome to inhibit expression of co-stimulatory molecules needed for T cell effector function... most acclaimed form of immunotherapy in recent times. These engineered T cells are capable of recognizing...immunotherapy targets can be utilized at the same time. A combination of tolerance marker blockade and ...