...targeted tdTomato and pre-synaptic targeted synaptophysin tagged eGFP. AAV phSyn1(S)-FLEX-tdTomato-T2A-SypEGFP-WPRE...deployed around the world to combat severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus...ready-to-use viral preps with more on the way. S5E2 regulatory element containing vectors developed in the Jordane...Flp-dependent DREADDs viral vectors. Both inhibitory and excitatory DREADDS from the Gether lab are now available...

...specific genes. Many years ago, people in the laboratory became frustrated with using glass cloning rings...sterile truncated pipet tips. Indeed, an experienced operator can pick 48 clones from a 10 cm dish in 15- 20...YouTube video. Before returning the plate to the incubator, additional growth medium is added to about halfway...two 48-well plates of clones) which enables investigators to identify specific clones with desirable properties...Iris Lindberg is a professor of Neurobiology and Anatomy at the University of Maryland Medical School in...

...is Head of Synthetic Biology at the Sainsbury Laboratory, where she often feels more like an engineer ...the toolkit. We have included a wider range of terminators than are commonly used as we have tested them...system. The paper also gives quantitative data for regulatory elements, which has not been available for plants...easy to express genes in, which makes it a good laboratory workhorse. Tips & tricks Addgene: Do you have...start. They might want to use our promoters and terminators, but then they aren’t sure how to convert their...

...in a year-long biochemistry laboratory : Part II – research-based laboratory – a semester-long research...confidence in their research capabilities and laboratory and critical thinking skills (Knutson et al.,...improvisation and the latter making him a more effective motivator (“be like Tony Robbins!” he encourages faculty...research-based learning in a year-long biochemistry laboratory: Part I – guided inquiry – purification and characterization...

... are successful base editors in rice, tomato, wheat, and potato. Target-AID and rAPOBEC based CBEs are... HDR methods for rice crop improvement in the laboratory of Yinong Yang at PSU.   References 1. Komor,...Zenpei, et al. "Targeted base editing in rice and tomato using a CRISPR-Cas9 cytidine deaminase fusion."...The CRISPR-Cas Revolution, Cold Spring Harbor Laboratory (2018). Additional resources on the Addgene blog...

...the cytoplasm. That is, unfortunately, some investigators realize too late that they have missed the impact...environment where it will localize [4]. Most investigators are surprised to learn that FPs are often poorly...overestimation of fusion protein activity. We encourage investigators to characterize FP-fusion proteins functionally...Einstein College of Medicine in the Department of Anatomy and Structural Biology. His interests include the...Medicine. Her doctoral research was conducted in the laboratory of Erik Lee Snapp and focused on optimizing fluorescent...

...upregulate an alternative pathway or induce other compensatory mechanisms. How can protein function be dissected...protein in Rosetta E. coli. New Lentiviral CRISPR activator and repressor plasmids from Scot Wolfe's lab. .... These include Tet-inducible CRISPR activators and repressor plasmids. (Kearns et al., Development. 2014...basic parts (promoters, UTRs, coding sequences, terminators, etc) into individual Level 0 plasmids, or choose...unit (Ex: a promoter, 5’UTR, coding region, and terminator). Next, up to six Level 1 modules can be similarly...in optimizing TALEN assembly and activity? The laboratory of Takashi Yamamoto has created a complete TALEN...of the gene of interest, C-terminal tag, plant terminator, and plant resistance cassette) in pUC19 based...

...blocker isolated from a designed repeat protein combinatorial library by in vivo functional screen." Scientific...production of a downstream peptide, the transcription terminators in this series are just the tool you need. The...this portion of the BIOFAB Plasmid Set contain terminators of varying strengths centered within GFP/RFP ...characterized in Cambray et al., 2013. These terminators can be used to precisely tune your polycistronic...Kit for some insights on its potential from its creators. We hope you’ll find these kits useful in your...

...list from your current P.I., lab manager, or administrator containing all of the materials that you’ve ...financial values. All too often, we see early investigators making unnecessary short term sacrifices that...solutions to reach your goals. For example, a junior investigator turned down a faculty position because none ...Happy Sciencing, Damien   Damien Wilpitz is a laboratory research management consultant and coach in Boston...research scientists create productive and manageable laboratory environments. His business skills enhance his...

...construct (about 8 in Brainbow-1.0 mice.) The combinatorial effect of multiple, independent recombination...woodchuck hepatitis virus post-transcriptional regulatory element (WPRE), commonly used to increase transgene...plasmids References: Transgenic strategies for combinatorial expression of fluorescent proteins in the nervous... Lgr5 stem cells. Snippert HJ, van der Flier LG, Sato T, van Es JH, van den Born M, Kroon-Veenboer C, ...PubMed. Multiplex cell and lineage tracking with combinatorial labels. Loulier K, Barry R, Mahou P, Le Franc...

... a variety of species, including the standard laboratory mouse. It’s also made targeted insertions relatively... Figure 6: Diagram of ovary and oviduct anatomy with the equipment used for i-GONAD. A micropipette...  Many thanks to Channabasavaiah B. Gurumurthy, Masato Ohtsuka, and Rolen M. Quadros for assistance with...Rolen M. Quadros, Channabasavaiah B Gurumurthy, and Masato Ohtsuka. “Easi-CRISPR for creating knock-in and...10.1038/nprot.2017.153 PubMed PMID: 29266098 Ohtsuka, Masato, et al. i-GONAD: a robust method for in situ germline...

...light within a specific range of wavelengths. Excitatory channels allow positively charged ions such as...combine excitation and inhibition by using both excitatory and inhibitory channels activated at different... the opsin.      Figure 2: Diagram of excitatory (ChR2) activation by light showing ions flowing... a single pathway to activate various protein regulators at different points in the pathway. Photoswitches...., Shane, J. C., McKnight, D. J., Yoshizawa, S., Kato, H. E., Ganguli, S., & Deisseroth, K. (2019). Cortical...

...in immune cells have identified many putative regulators of immunity. However, in vitro screens cannot...responses to cancer. The screen recovered known regulators, such as Ptpn2, Pdcd1, and Zc3h12a, demonstrating...Moreover, the screen identified putative novel regulators of CD8+ T cell responses in tumors, demonstrating...LaFleur is a Postdoctoral Fellow in Arlene Sharpe’s laboratory at Harvard Medical School and is interested in...

...establish your own lab and get an independent investigator grant (e.g. an R01 from the NIH, in the United...can be found during conversations with senior investigators. Discussions can reveal mutual scientific interests...makes time for you, as needed, this is a strong indicator that this person cares about your science and ...manuscript reviews, etc. You'll also meet future collaborators and get access to unpublished information and...Einstein College of Medicine in the Department of Anatomy and Structural Biology, and now serves as the Director...

...customized to control transgene expression. Gene regulatory elements [promoters and enhancers, miRNA target...Abbreviations: AAV, adeno-associated virus; GRE, gene regulatory element; NHP, non-human primate. Many of...broad):  enhanced coverage for inhibitory versus excitatory neurons Mice, rats Yes (LY6A) Chan et al...AAV.CAP-B10 CNS: balanced coverage of inhibitory and excitatory neurons Mice, marmosets Unknown Brown et... frequently been used for screening candidate regulatory elements such as enhancers and promoters for ... its tropism is relatively more biased toward excitatory neurons than AAV-PHP.eB (Fig. 4c, Jang et al....., … Tasic, B. (2021). Enhancer viruses for combinatorial cell-subclass-specific labeling. Neuron,109(...

... availability of this strain from The Jackson Laboratory. Originally developed and characterized by Stanley...mouse strain is regaining popularity. The Jackson Laboratory, a nonprofit research institution, has been able...clinical manifestations, transmissibility, and laboratory studies of COVID-19: an observational study. ...of K18-hACE2 Mice Infected with Severe Acute Respiratory Syndrome Coronavirus. JVI 81:813–821 . https:...

...technology (Yesbolatova et al., 2019). Find the mAID tagging toolkit at Addgene Yesbolatova et al., Methods...tagging of proteins with mAID and other tags (Yesbolatova et al., 2019) using CRISPR. This toolkit makes...neurons. Earlier this year, the Hegemann lab and collaborators report their discovery of a new family of seven...

...immunosurveillance. The presented study analysed a model of hepatocellular carcinoma, a cancer which can develop from ...self-limiting effects kick in trying to resolve the inflammatory state by inducing immunosuppressive measures...Chimeric Antigen Receptors) CAR T cells and PD-1 modulators. An exciting development in immunotherapy is ...a researcher to control immune signaling and inflammatory responses remotely with spatiotemporal precision...

...heterologous envelope or a post-transcriptional regulatory element can be used for either type of retrovirus...clinical settings. AAV is also a cool tool in the laboratory! It is ideal for whole animal administration,...population. Some adenoviruses are known to cause respiratory disease across species, as well as cancer in ...strains have been engineered to induce a lower inflammatory response within hosts. Relative to the other...

...activation of keratinocytes, which release inflammatory mediators that trigger the activation of dendritic...Generalized pustular psoriasis is a chronic autoinflammatory skin disease estimated to affect 0.27 to 4.6...IL-36 cytokines are a potent inducer of an autoinflammatory loop: keratinocytes both respond to and release...

...CRISPR for that, too! CRISPR activators (CRISPRa) are transcriptional activators fused to a dead Cas9. The...increase expression. There are several flavors of activators which can be used with this approach - read more...them on our CRISPRa blog! Fig. 1 CRISPR activator and repressor mechanisms Want to transiently...

...sword – balancing antiviral benefits with the inflammatory response associated with the introduction of...threshold and thus do not typically activate an inflammatory response. However, if dsRNA sensors are mutated...its way outside of the mitochondria, the same inflammatory response can activate. These types of cellular... disorders, including some forms of lupus and rheumatoid arthritis. These types of immune responses are...

...available to counteract the toxin. If, however, an investigator were to successfully clone an insert into the...insertion will completely replace ccdB with the investigator’s insert of interest. Hence correct clones are... behind the “swapping out” of ccdB with the investigator’s gene of interest.   CcdB resistant E. ...for non-cytotoxic proteins which retain their regulatory functions.” Mol Microbiol. 1995 Mar;15(6):1031...

...used red fluorescent proteins dsRed, mCherry, and TdTomato do not induce transcription. Thus, T-DDOGs can...expression robustly activated the reporter gene TdTomato, whose expression was absent without electroporated...T-DDOGs with two mouse GFP reporter lines; again, TdTomato was seen only in cells with GFP, at a high activation...frequency of 56-93%. In the converse test, 98% of TdTomato expressing cells were GFP+, indicating a robust...

... infections along with a mild or exacerbated inflammatory response which leads to nearly 25% child mortality...understood.  In line with this, Prof. Videira and collaborators will use Addgene plasmids to immortalize CDG...discovery led them to investigate the use of a common laboratory chemical, guanidine hydrochloride, as a potential... 1939 for the treatment of botulism and Lambert-Eaton myasthenic syndrome (LEMS) because, at high concentrations...

...bone marrow (BM) cells, primary human hepatocytes, and hematopoietic stem cells (HSCs) using Nanoblades....deliver Cas9 variants, such as a transcriptional activator dCas9-VPR, if these variants are fused to the ...from GFP transgenic mouse  GFP  ~60-65% Human hepatocytes  Myd88  ~50% Human HSCs  Myd88  ~50% Table...

..., 50ms “off”.  A pulse generator such as the A.M.P.I Master-9 Pulse Stimulator, which can control output...cable is threaded through—and fixed to—an optical insulator called a ferrule (below). The fiber optic cable...to give the animal the full range of motion, a commutator allows the patch cable to connect to the laser...

... bank account, business cards, incorporation, laboratory lease, etc. I still flip to that page in my lab...business. We ended up applying to a business accelerator freely stating that we had an idea, knew how ...rough day, and we have great partners in this accelerator program who want us to succeed. We’re excited...duty to pursue a degree in a synthetic biology laboratory and received her M.S. from CSU in 2014. Outside...

...common use of intradermal injections is studying inflammatory responses, as this method is useful for analyzing... correct location requires a lot of skill and anatomical knowledge. Intrathecal injections are used for...Routes of administration. In H. Hedrich (Ed.), The Laboratory Mouse. Academic Press. Turner, P. V., Brabb, ...M. A. (2011). Administration of substances to laboratory animals: routes of administration and factors...

...with Addgene by year. (C) Cumulative number of laboratories that have deposited CRISPR plasmids by year....nuclease into three functional groups: nucleases, modulators, and chimeric effectors. Nucleases generate a... a classic double-stranded break, while modulators regulate transcription and chimeric efforts make a ...deposited and distributed CRISPR plasmids. But modulators and chimeric effectors, made available in 2013...

...up with poor droplet quality. Using a droplet generator, the sample and the droplet generation oil are...Pipette slowly If you are using a manual droplet generator, you will have to be careful about transferring...transferring your droplets from the droplet generator to the PCR plate. Maintaining a high droplet count is important...sample. Even if you are using an automated droplet generator, you should avoid pipetting too quickly when making...

...m a father of three, a scientist and science communicator, and I strive to be a volunteer and contribute...scientist. When our elementary school needed a new coordinator for the annual Math & Science Night, I took the...contacting our school district’s science curriculum coordinator with minimal luck, but eventually the 8th grade...great feedback from teachers and other science communicators.  What can CRISPR do? I spent more time on...

... J. (2002) Arabidopsis : a laboratory manual. Cold Spring Harbor Laboratory Press. Available at: https...insertion likely precipitated domestication of sweet potatoes (Kyndt et al., 2015)! It sounds simple, and it.... et al. (2015) ‘The genome of cultivated sweet potato contains Agrobacterium T-DNAs with expressed genes...

...is the Laboratory Operations Manager at Addgene.  In addition to helping with daily laboratory operations...through barcoded tubes that are scanned into our Laboratory Information Management System (LIMS), a database...operations, she also works on improving laboratory efficiency through automation.       More Resource Avaliable...

...amount of grant funds available and the increasing regulatory process for grants has fostered the demand for...professional. Dr. Cook served as a principal investigator for several research grants. One of the benefits...benefits of being a principal investigator is that you are asked to review grant proposals from others. ...worked for a health department and then for the regulatory services department as our healthy homes program...

...time-consuming, and complex to use outside of standard laboratories. In the case of the COVID-19 pandemic, qPCR ...platforms to reliably detect nucleic acids at the atomolar level. The Zhang lab has adapted natural RNase...reporter enables SHERLOCK to detect DNA or RNA with atomolar sensitivity and single base pair mismatch specificity... of HPV, HPV16 and HPV18, from human cells, at atomolar levels, within one hour. DETECTR thus has the ...

...recently was invited to attend the first Principal Investigator meeting of the NSF EDGE (Enabling Discovery ...parenting strategies and make important ecological indicators since species live in water and on land. Current...that tastes yucky (and is poisonous) for bird predators - birds eat a caterpillar once, vomit, and never... expensive. They are using a cool fluorescent indicator system to rapidly identify recombinants. Arthropods...

...simple as a piece of paper on the front of the incubator containing the passage number, cell line, and ...book and on the actual flasks of cells in the incubator). This means any other scientist who wishes to...The LGC Group also provides standards and has laboratories to carry our independent quality control for...culture contamination." Archives of pathology & laboratory medicine 133.9 (2009): 1463-1467. PubMed PMID...

... vector preps expressing the Voltron2 voltage indicator, created by the Schreiter Lab and the GENIE Project... available! Building on their earlier Voltron indicators (a fusion of Ace2N and HaloTag, Figure 4), they...The same A-to-D mutation in the related voltage indicator Ace2N-mNeon also led to similar improvements in...fluorescent signal at baseline voltage), the investigators found that administering these AAVs with a dilute...

...characteristics that make them ideal for studying in a laboratory environment. We’ve covered five model organisms...have been specifically developed to thrive in laboratory settings. One of its most attractive features...characterized genome, and is easily maintained in a laboratory environment. What sets Arabidopsis apart from.... The Norway rat, from an obnoxious pest to a laboratory pet. ELife, 9. https://doi.org/10.7554/elife.50651...