We narrowed to 1,029 results for: ELL
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TypeBlog Post...would rather fund excellent people with mediocre ideas than mediocre people with excellent ideas. I am lucky...your life making connections and finding mentors -- well, it's time you call upon these people for guidance...Don’t go it alone. You’ll find that no matter how well you know your idea, you will rarely succeed in launching...partners. You need a dedicated team that you know well and trust. This will allow you to discuss each ...two close friends, Sreyan Chowdhury and Clinton Buell, who have very diverse educational backgrounds (...
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Advice for Moving Into Sales After Your Science Postdoc
TypeBlog Post...Fhearraigh, PhD, of www.phdadvice.com. I sold out, well in part that’s what leaving academia feels like ...would continue on with my academic dream of studying cell division. However, after 6 years in the business...products, so you have some of the best ideas on how to sell these products to other scientists. Tailoring my...introduction to the financial world. It teaches you about selling products which is an important part of the business...Cambridge University where he studied mechanisms of cell division. Recently Seán got the entrepreneurial ...information on ELISA troubleshooting, ELISA kits and cell death assays. You can find Seán on twitter @Macfhearraigh... -
Interview: Ed Boyden on Optogenetics, Neuroscience, and the Future of Neuroengineering
TypeBlog Post...an awake monkey is different from a cell in a dish. I always tell people, 'Be driven by your science.'...for neuroscience back in 2005, both activators as well as silencers of neural activity. Since then, there...to Addgene in a matter of days. Addgene: Can you tell us a little more about this new two-color tool and...and see how different parts of the brain or the cells within them work together or perhaps compete against...two-photon properties, so you could activate single cells in arbitrary patterns in the brain. Addgene: Do ... -
The Crispr Babies Saga Shows the Need for Action, Not More Delays
TypeBlog Post...now. Many thanks to our guest blogger, Kelly Hills! Kelly Hills is a founding bioethicist of the consulting...This post was contributed by Kelly Hills, founding bioethicist of Rogue Bioethics. On November 25, the... the MIT Technology Review dropped a bombshell report. A scientist working in China was using CRISPR/Cas9...separate things happened nearly simultaneously: The Wellcome Trust funded a center for mitochondrial research...consulting firm Rogue Bioethics. Kelly utilizes her firsthand experience in the software industry to inform... -
Plasmids 101: Gram Negative and Positive Bacteria
TypeBlog Post...negative bacteria is due to differences in cell wall composition. Cell wall structure Gram positive bacteria...used in the staining process bleached another, Klebsiella pneumoniae. After realizing this was a repeatable...membrane. Common gram negative bacteria include salmonella and E. coli. Because of their thicker peptidoglycan...complex within the peptidoglycan layer, staining the cell purple. When the alcohol is added, it dehydrates...Silhavy, TJ., Kahne, D. & Walker S. The Bacterial Cell Envelope. Cold Spring Harb Perspect Biol. 2(5) a000414... -
DAP arrays for multiplex gene editing
TypeBlog Post...different types of human cells, including T cells and hematopoietic stem and progenitor cells. Now with the DAP...to enable multiplex gene editing in a variety of cells and organisms. However, lengthy genetic payloads... over 50% averaging editing efficiency in human cells. MBE with DAP array also exhibited reduced Cas9-... editing, and nuclease editing, in virtually any cell type or organisms of interest with only minimal ... -
Revamp Your Lab Meetings With Creative Virtual Collaboration
TypeBlog Post...blogger Matteo Tardelli! Matteo Tardelli, PhD is Postdoctoral Scientist at Weill Cornell Medicine in NYC...was contributed by Matteo Tardelli, a postdoctoral scientist at Weill Cornell Medicine. Discussing data...Hamilton DP, Denfeld BA, de Eyto E, Hampton SE, Keller PS, Sharma S, Lewis ASL, Weyhenmeyer GA, O’Reilly... -
Addgene-Seeding Labs plasmid grant round 3: microbial drug resistance and biofilms, antidiabetic plant components, visualizing brain injury - and a glowing robot fish
TypeBlog Post...multispecies biofilm formed by Pseudomonas spp. and Klebsiella spp. Dr. Devarshi Gajjar, The M. S. University...research focuses on a collection of pathogenic Klebsiella and Pseudomonas isolates causing urinary tract...unknown regulatory regions in Pseudomonas and Klebsiella spp. The effects of these deletions on biofilm...major pathophysiological complications of diabetes mellitus. Recombinant human proteins that play a role in... Researcher who investigates the molecular and cellular etiopathomechanisms of ischemic stroke, the most... reduced blood flow. Novel treatment options, as well as non-pharmacological solutions to modify the course... -
With Our New Viral Service, We're Taking CRISPR Further!
TypeBlog Post...of viruses used to deliver CRISPR components to cells or organisms under study. In the past we’ve left... they want to do is to quickly generate a single cell line and proceed to use it for experiments.” - ...screen for genes involved in a wide variety of cellular processes. While these are widely requested and...representation in the viral prep, infect target cells, and, finally, conduct the screen. To help alleviate...of this labor for our users, we now provide the Brunello (targeting human genes) and Brie (targeting mouse...deliver gRNA and therefore, must be used to infect a cell line already expressing Cas9. As John Doench (Broad... -
Optimizing Donor DNA for Enhanced CRISPR Genome Editing
TypeBlog Post... between Cas9 (which causes the breaks) and host cell DNA repair factors (which repair those breaks) makes...example, correcting the allele that causes sickle cell anemia. However, HDR occurs much less frequently...a structure could drive sequence replacement in cells. Targeting three dCas9 molecules to a precisely ...functional Cas9 protein and the targeting guide RNA into cells by electroporation. Results from our lab and others... HDR. The detailed protocol for nucleofection as well as the sequences of our validated editing reagents...Highly efficient RNA-guided genome editing in human cells via delivery of purified Cas9 ribonucleoproteins... -
AAV Q&A with Tim Miles
TypeBlog Post...address all your unanswered questions via text! (well, maybe not all of them. We still haven’t gotten ...and would like to get it working in microglia as well. A: We are not aware of any available AAV for efficiently...co-delivering different AAV particles into the same cells in vivo? A: This will be highly dependent on vector...particularly one that can target the epithelial cells in the blood-brain barrier? A: For liver, we recommend...worth investigating for both Q6 & Q7.For epithelial cells of the blood brain barrier we would currently recommend...and to all our participants for their time and excellent questions. Additional Resources on the Addgene... -
Viral Vectors 101: The AAV Retrograde Serotype
TypeBlog Post... into the cell. This means that not all neurons and pathways will be transduced equally well by AAV-retrograde...they either trace downstream from the soma (neuron cell body) toward the axon (anterograde tracer) or upstream...them useful tracers, but finding a virus that works well for retrograde tracing can be difficult. The in ... Data Hub!) The cortical pathways to the IC were well transduced by AAVrg while other pathways such as...chemogenetic vector directly into the region where the cell bodies of the projection neurons reside. Impact... -
Developing a cold tolerant E. coli using specialized chaperones
TypeBlog Post...government agencies, active researchers, as well as fellow iGEM teams. We were the proud hosts for the...industrial or research purposes requires stable host cells (the simplest example would be a transformed mesophilic...successful) as host systems since their biology is well studied, and they are convenient to maintain under..., since mesophilic chaperones would not function well under suboptimal conditions. This means recombinant...CRISPR-Cas mediated editing for psychrophilic systems as well as engineering and regulating psychrophilic recombinant... -
REPLACR Mutagenesis: Replacing In Vitro Recombination Methods
TypeBlog Post...electrocompetent cells in which the phage recombination machinery is induced using L-arabinose. These cells can be...function of certain regions in a promoter or gene, as well as to study the effects of inactivating/activating...coli to undergo nick repair. This method can work well, but poor template digestion can result in a high...required in recombineering. REPLACR-mutagenesis is also well suited for insertions and deletions. The figure ...Enzymes Required No No Yes Yes Lab Made Competent Cells Required Yes No No No Comparison of REPLACR-mutagenesis... -
Protein Tagging with CRISPR/Cas9: A Conversation with Mendenhall and Myers
TypeBlog Post...also gives cells antibiotic resistance to neomycin so that you can select the population of cells with the...transcription factor. The FLAG antibody has been well characterized and the system allows you to add this...Only a small percentage of the proteins inside a cell have a commercial antibody against them. There’s...antibody. It’s an epitope not found in human or other cells. It was made artificially. That means you aren’t...lab already should be able to run this protocol as well. We would also like to put some social pressure ... -
Troubleshooting Your Plasmid Cloning Experiment
TypeBlog Post...product, the transformation may not result in any cells, or all the colonies screened might not contain ...i.e. primer3) or provided by the major companies selling PCR reagents. Nowadays commercial polymerases are...verify the strain and the efficacy of your competent cells (when they get old, they lose competency). If all... look at your experimental design. Full lawn of cells. The antibiotic in your plate might not work. Check... the incubator. The small colonies are called satellite colonies and they form when all the antibiotic...acknowledged in hundreds of publications including Cell, Nature, and other journals. Paolo Colombi is ... -
The time and cost required to make a plasmid
TypeBlog Post...we virtually always use every single well in a 96-well (or 384-well) plate! How much do repositories help...in this post is a mathematical story well beyond my ability to tell. But even from these rough estimates...depositors have helped us save the scientific community well over $80 million on plasmids alone. (And we expect... -
Tagging Optogenetics and Chemogenetics Receptors: Fluorescent Proteins and Other Options
TypeBlog Post... be expressed in the same cell, but not necessarily at the same subcellular compartment as the opsin or...modulate the activity of neurons and other brain cells. Since the opsins or chemogenetic receptors used...neuropil conceals identification of opsin-expressing cells). For other things to keep in mind when making fluorescent...cerevisiae by use of an epitope addition method. Mol Cell Biol 8:2159–2165 . https://doi.org/10.1128/mcb.8.5.2159...Paré J, Bonaventura J, Richie CT, Michaelides M, Mueller SAL, Roseboom PH, Oler JA, Kalin NH, Hall RA, Smith... -
Enhancing CRISPR Targeting Specificity with eSpCas9, SpCas9-HF1, & HypaCas9
TypeBlog Post...-HF1, & HypaCas9. The off-target problem It is well known that CRISPR/Cas9 genome editing can result... the EMX1 locus in human embryonic kidney (HEK) cells, 11 of these initial mutants retained the on-target...off-target activity at multiple genomic locations in HEK cells. Further testing of two of the mutants, SpCas9(K855A... showed that these mutants are not toxic to HEK cells and that similar mutations can improve the specificity... researchers to make precise edits in mammalian cells and may decrease worries about off target effects... Cas9 in complex with guide RNA and target DNA." Cell 156.5 (2014): 935-949. PubMed PMID: 24529477. PubMed... -
Plasmids 101: In-house NGS Plasmid Quality Control Now at Addgene
TypeBlog Post...process will begin. Partnering with seqWell, we are using the plexWell technology to easily and quickly ...ever worked to sequence DNA on NGS platforms will tell you, while the process has become much more refined...assembly process. Again, thanks to our partners at seqWell we make use of a pipeline that takes the raw data...necessary adapter sequences for Illumina sequencing as well as unique barcodes per sample so we can assign sequencing...Tasissa M, Kostman M, Tillgren M, Makana Hanley L, Mueller I, Mitsopoulos A, Fan M (2020) A Novel Next-Generation... -
Savvy Advocates Needed to Navigate a Scientific Enterprise in Flux
TypeBlog Post...Communications, Harvard University Ray Howell -Founder of Howell Communications Philip Brenner, PhD - Assistant...scientific enterprise and the way it is funded have been well documented over the past 2 years (Alberts et al,... to shape the sector throughout their careers (McDowell et al, F1000Research, 2014; Dolan, F100Research...relationship, along with, of course, union dues. Time will tell the extent of the implications of this ruling and...27175853. Pubmed Central PMCID: PMC4866822. 3. McDowell, Gary S., et al. "Shaping the Future of Research... -
Mobile-CRISPRi: Bringing CRISPRi to Diverse Bacteria
TypeBlog Post... acid (DAP), a component of the cell wall. Once in the recipient cell, the CRISPRi DNA (purple) flanked... is propagated during normal DNA replication and cell division. Mobile-CRISPRi does not disrupt gene function...mobilize these plasmids for transfer to recipient cells during a tri-parental mating scheme. Mobile-CRISPRi...bacteria associated with human disease (S. aureus, Klebsiella pneumoniae, and P. aeruginosa) in addition to...antibiotic trimethoprim. When Enterobacter aerogenes, Klebsiella pneumoniae, and P. aeruginosa (all opportunistic... -
CrispyCrunch: High-throughput Design and Analysis of CRISPR+HDR Experiments
TypeBlog Post...AmpliCan We chose to adapt Crispresso by Luca Pinello and Kendell Clement for batch analysis because of its...Leonetti Ryan Leenay Jason Li Max Haeussler Luca Pinello Kendell Clement Andy May Additional resources on the...members have taken up to a month to design a single 96-well plate for HDR that satisfy all requirements. To ...all the standard information that goes into a 96-well plate and comes out of it. Biotech companies are... -
Antibodies 101: Flow Compensation
TypeBlog Post... make it difficult to properly gate your desired cell population. By compensating between fluorescent ...online manual for your cytometer should be able to tell you the laser configuration. In general, if two ...Figure 1A). Applying compensation means you are telling your cytometer to adjust the fluorescent signals... your fluorescent markers are staining for. If a cell is positive for both markers, it will show up as...Figure 3: Flow plot for a double positive sample. The cell population in the top right corner of the plot is...learning! Ashlyn has a research background in CD8+ T cells and cancer immunotherapies. She is currently a research... -
Degrading DNA with Cascade-Cas3
TypeBlog Post...there isn’t a template strand left behind that the cell can use to repair. Figure 3: General overview...Occasional cleaving of the target strand is not as well understood but likely occurs in similar regions....complex in nucleotide unwinding steps. Molecular Cell, 70(3), 385-394.e3. https://doi.org/10.1016/j.molcel...formation by a type I-C CRISPR Cascade. Molecular Cell, 83(5), 746-758.e5. https://doi.org/10.1016/j.molcel...broad and unidirectional genome editing in human cells. Nature Communications, 10(1). https://doi.org/10.1038...CRISPR–Cas systems for genome engineering in human cells. Nature Biotechnology, 37(12), 1471–1477. https:... -
Anatomy of a Plasmid Page at Addgene
TypeBlog Post...different kinds of information on our plasmid pages? Well, to help make the most of these pages, we've written...Addgene’s plasmids is an indication of popularity. A yellow flame indicates that a plasmid has been ordered...process and these can be found in this location as well. These maps make it easy for scientists to see if...analyzed using Addgene’s Analyze Sequence tool. Excellent, how can I check if the plasmid will be within...resistance, and optimal growth temperature are listed, as well as the name of the strain. Most Addgene plasmids...such as some of our viral vectors, in NEB Stable cells. Resource information: This section contains information... -
Interview: Nicola Patron on Plant Synthetic Biology, MoClo, and More
TypeBlog Post...favorite parts for plant synthetic biology Addgene: Tell me about the Golden Gate Modular Cloning (MoClo)...really easy to clone genes for expression in plant cells. Sylvester and I chose the 96 most useful things...used as we have tested them to show that they work well in plants and give similar stability to transcripts... parts available in your lab - if you get the 96-well kit - everything is so easy. We have undergrads ...a real person, so that students see academics as well-rounded human beings. It’s also a great communication... that want to have a big impact on the world and tell them to work in plant science. I think their chances... -
Interview: Hodaka Fujii on enChIP, New CRISPR Tools, and More
TypeBlog Post...CRISPR complex much easier in various cells, including primary mouse cells. We believe that this system will...In 2014, you joined Addgene's Advisory Board, as well as the Editorial Board of Scientific Reports and...US, I very much enjoyed interactions with many excellent colleagues at NYU and visiting scientists, such...In addition, I've been very lucky to work with excellent colleagues in my lab. On the other hand, I feel...good for Japanese science. I really miss the intellectual environment in the US in this regard. In addition... -
Plasmids 101: Gateway Cloning
TypeBlog Post...choose a molecular cloning technique that will work well with a given set of resources, time, and experimental...the next step is to transform competent E. coli cells and select the positive clones. The entry clone ... expression of the human KRAS gene in mammalian cells. STEP 1: Generate an Entry Clone There are a few...ccdB gene. Since the ccdB product is toxic to the cell, Gateway cloning efficiency can reach >99%. ...digest! Then, you can transform or transfect the cells that you want to use for your experiments and verify...Phage (Lambda) and Higher Organisms (Cambridge, MA: Cell Press). Additional Resources on the Addgene Blog... -
Lab to Office Culture Shock
TypeBlog Post... it, and many of my scientist colleagues have as well. Making the transition from the bench can be jarring...scientist going through even the worst experimental dry spells. Losing that sense of discovery can be hard since...place. The feeling of camaraderie one gets from a well-functioning lab team. Obviously not everyone gets...applying. I probably did not respond to her question as well then as I would now. I would have to repress laughter...the right set of tools to help you do your job as well as possible. It’s in their best interest to do so...the newer members of our Scientist team says, “a well-run meeting where everyone has a voice and the group... -
Harnessing TnpB for Genome Editing: A Compact and Versatile Tool for Your Research
TypeBlog Post...editing reagents into cells. Viral vectors, because of their natural ability to infect cells, have emerged as...biologist, academic, and researcher. Among the most well-known CRISPR-associated nucleases are Cas9 and Cas12a...which poses a challenge for efficient delivery into cells, particularly via viral vectors. While viral delivery...for gene editing and allows targeting of specific cell types, it has a limited capacity to carry large ...biolistic, or PEG-mediated transformation into plant cells. Conclusion & Future Directions TnpB represents... -
Tips and Tricks for Using Golden Gate Modular Cloning (MoClo)
TypeBlog Post...recommend transforming into chemically competent cells in a 96-well PCR plate. Always confirm your results Congratulations... sequences of the parts and acceptor vectors, as well as the restriction enzymes, and it will create the...transfer and the restriction–ligation reactions in a 96-well (or higher!) format. Liquid handling robots are ...Dr. Rob Hurt for high-throughput minipreps in 96-well plates. Plate wisely You will likely see fewer numbers... this, you can increase the volume of competent cells you plate after each transformation and/or use different... -
CRISPR 101: Any Base Transversion Editors
TypeBlog Post... edit A → Y (Y = T or C) in 67.17% of screening cells (Tong, Wang, et al., 2023). Based on this promising...the base editor. Black arrows indicate natural cellular processes. Created with BioRender.com. The...Cas9 nicks the non-edited strand to encourage the cell to repair the DNA using the edited strand as a template... efficient A•T-to-C•G base editing in mammalian cells and embryos. Nature Biotechnology, 42(4), 638–650...programmable T-to-G and T-to-C base editing. Molecular Cell, 84(7), 1257-1270.e6. https://doi.org/10.1016/j....efficient T-to-G and C-to-G editing in mammalian cells. Nature Biotechnology, 42(10), 1538–1547. https:... -
Hot Plasmids: Fall 2024
TypeBlog Post...(or even in the middle of proteins) and performs well in many different antibody-based applications. ...Randall, R. E., Young, D. F., Goswami, K. K., & Russell, W. C. (1987). Isolation and characterization of...useful addition to the toolkit for visualizing cellular processes in C. elegans: mScarlet3 for applications...biologically relevant forces at focal adhesions in living cells in several different contexts. Figure ...substrate furimazine. Right: Demonstration of PILATeS. Cell adhesion to a PILATeS-functionalized coverslip creates...minutes. Find PILATeS plasmids here! Zhong, B. L., Elliot, J. M., Wang, P., Li, H., Hall, R. N., Wang, B.... -
Binning Singletons: Tackling Conference Networking When You Don’t Know Anyone
TypeBlog Post...occurs when bacterial cells regulate gene expression in response to changes in cell density. At the mixer...cohort of friends and colleagues had moved on as well, so I knew very few people. I didn’t really fit ...Singletons gained from spending time with a mentor, as well as having a cohort experiencing similar situations...introduced to several people by their mentor, as well as their bin-mates. They liked us. They really ...Hendrickson, Jonathan Jacobs, Mo Kaze, Christina Kellogg, Heather Martinez, Pat Schloss, Tara Smith, Marc... -
Top Requested Plasmid of 2017 - pMD2.G
TypeBlog Post...used to deliver Cas9 and other genes to mammalian cells. For this reason, the top requested plasmid of 2017... coat the virus and help determine the types of cells that it can infect. The VSV-G envelope is widely... lentiviruses to infect many different types of cells. This means researchers who use VSV-G can deliver...deliver their genes of interest to a variety of cells and study their effects. To date, nearly 900 articles...in studies of the secretome, ubiquitin ligase, cellular senescence, autophagy, CRISPR, and so much more... -
An Interview with Connie Cepko - Gene Therapy, Plasmid Tools and Insights to Success
TypeBlog Post...fluorescent proteins as scaffolds for cell-specific gene manipulation.” Cell 154(4) (2013): 928-939. PubMed ...Cepko Interested in Working on the Eye 9:05-10:16 : Cell Targeting in Retinal Gene Therapies 10:17-14:34 ...28:39 : Methods for Delivering Multiple Genes to Cells for Gene Therapy 28:40-35:10 : GFP Activatable Tools...publication at Addgene. 2. Tang, Jonathan C.Y., et al. “Cell type-specific manipulation with GFP-dependent Cre... -
Transferable Skills Guide: Conflict Resolution
TypeBlog Post...other people, but resolving your own conflicts as well. What I really brought to the table was empathy,...you can identify them in yourself and learn how to sell them. Types of Conflict Before explaining my approach...occurs when people or teams who normally get along well with each other disagree about a specific decision...good listener absorbs and remembers and makes the teller feel heard. I am not always a stupendous listener...will accept. If you are able to broker a compromise well, you can help everyone walk away from a conflict...solution C. And if a conflict resolver does their job well, not only will solution C be better than either ... -
CRISPR 101: Homology Directed Repair
TypeBlog Post...double-strand break (DSB). DSBs can be caused by intracellular factors such as nucleases and reactive oxygen...repair fix DNA double-strand breaks? In eukaryotic cells, repair of DSBs occurs primarily by two pathways...used, however, these are generally more toxic to cells than their plasmid counterparts. Another consideration...how to fix a broken relationship. Pardo B, et al. Cell Mol Life Sci (2009). PubMed. Regulation of DNA...break repair pathway choice. Shrivastav M, et al Cell Research (2008). PubMed. Mechanisms of DNA double-strand...Improve HDR Efficiency for CRISPR Editing in Human Cells Resources on Addgene.org Browse All CRISPR Plasmids... -
10 Steps to a Perfect Science Talk
TypeBlog Post...giving science talks and how to make them serve you well. The happy byproduct might just be a career opportunity...Listen to their stories and narratives (see below “Tell a story”) and note how they build suspense. Notice...host told me and wished I had been more careful. 3. Tell a story We are biologically programmed to listen...video for a job talk, but if you can get it working well, everybody loves a great animation. 8. Predict questions...good topic or story is all you need, if you do it well. Resist the urge to show all or even a lot of data... I hate it when I watch myself speak, but others tell me that I keep the talk moving enough that they ...