...gRNAs By Species Mammalian Bacteria Drosophila Plant C. elegans Yeast Zebrafish Xenopus CRISPR Resources ...transcription Virmilion none, need Cas9 plasmid Bullock and Port 49411 pCFD4-U6:1_U6:3tandemgRNAs dU6:1 and dU6:3...transcription Virmilion none, need Cas9 plasmid Bullock and Port 49330 pAc-sgRNA-Cas9 dU6 BspQI Transfection Puromycin...transcription Virmilion none, need Cas9 plasmid Bullock and Port 49409 pCFD2-dU6:2gRNA dU6:2 BbsI Injection or in...transcription Virmilion none, need Cas9 plasmid Bullock and Port 51026 U6-BbsI-crRNA dU6 BbsI Transfection none, ...

...of our most popular mammalian expression vectors, c-Flag pcDNA3, comes from the pcDNA series. It has the...consider as you make your choice. The first and most important thing you need to know is your expression system...have to make sure that your plasmid has been incorporated into the host organism, usually achieved with...basal expression level, induce expression at an opportune time or go all out with constitutive high level...Some of the more popular studies involve using a reporter vector to tag YGOI with lacZ and/or GFP (e.g. ...animals, gene targeting and genome modifications to reporter vectors (e.g. pBV-Luc) and gene silencing (e.g...

...Boeckman JX, Manning CF, Nguyen JT, Bechtold H, Dumitras C, Gong B, Nguyen K, van der List D, Murray KD, Engebrecht...and have always been, a community resource that supports both toolmakers and tool users. We encourage you...Educational Resources As part of our commitment to supporting your research, Addgene is creating an array of...Through our incredible community, including your support, we have enabled scientists to easily share plasmids...Studies Hybridoma Bank, a nonprofit that has been supporting hybridoma sharing within the community for decades...Immunofluorescence Addgene’s Antibody Guide The project is supported by a NIH BRAIN Initiative grant (U24NS119916 )...

... Deverman BE, Matho KS, Cetin A, Woodard K, Cepko C, Guerin KI, Rego MA, Ersing I, Bachle SM, Kamens J...’s ~4.4 kb or smaller. Some examples include a reporter like GFP, some smaller CRISPR-Cas proteins such...the Cre recombinase, are used to spatially and temporally control transgene expression. Lox sites are directional...sites lets researchers control the spatial and temporal activation of a transgene. Figure 2: FLEX...structure when transcribed which aids in nuclear export of mRNA and helps increase the expression of your...polyadenylation or polyA signal aids in the nuclear export of RNA and RNA translation, and promotes RNA transcript...

...∆G, however, can not be conjugated to PE due to a C-terminal glycine deletion, so it remains in the cytosol... development of several fluorescent autophagy reporters now allows for the measurement of autophagic flux...makes Keima useful for monitoring mitophagy. It’s important to remember that because Keima’s fluorescent relies... al. "Rosella: a fluorescent pH-biosensor for reporting vacuolar turnover of cytosol and organelles in...using a single fluorescent biosensor." Scientific reports 5 (2015): 12397. PubMed PMID: 26215030. PubMed ...

...Twitch calcium indicator, based on C-terminal domain of troponin C Griesbeck Lab Twitch Plasmids Oliver...GMP) FlincG3 (GFP-based cGMP sensor) for imaging in C. elegans neurons Using a Robust and Sensitive GFP-...Asymmetrical distribution of the second messenger c-di-GMP upon bacterial cell division. Science. 2010...Plasmids Joel Kralj Voltage Voltage sensors for use in C. elegans (Arch(D95N), MacQ-mCitrine, QuasAr and QuasAr-mOrange...cyclic AMP) Signaling reporter island (SiRI) with GFP-based fluorescent reporter cAMPr Spatial Multiplexing...maturation reporter (pmRFP-LC3) Dissection of the autophagosome maturation process by a novel reporter protein... DsRed1-E5 as a reporter for mitochondrial oxidative stress A novel MitoTimer reporter gene for mitochondrial...

... antibodies aren’t free to bind endogenous IgGs. (C) A third solution is to use Fab fragments against ...primary antibody that is directly conjugated to your reporter circumvents the issue. However, directly conjugated...which uses just a primary antibody that has your reporter (in this case a fluorophore) directly attached... as effective for other techniques, but it is important to be aware that the issue is not unique to IHC...

...) antibody-dependent cellular cytotoxicity (ADCC) c) antibody-dependent cellular phagocytosis (ADCP). ...starting a cascade of events that results in the pores being opened in the pathogen’s cell membrane (Dunkelberger... when holes are punched in their membranes, the pore'd pathogens quickly lyse and die.  Antibody dependent...

...released from nuclei by a short incubation at 37 °C. These short DNA fragments can then be purified for...MNase to antibody bound proteins (Figure 1). Importantly, since Protein A has weaker affinity for mouse... rather than to rely on nuclei extraction. They report that this change offers a significant improvement...

...under the United States Internal Revenue Code 501(c)(3). That means we were formed to benefit the public...do. From all of us to all of you. Thanks for supporting nonprofit, open science and Addgene in our mission...Collaboration with Fellow Nonprofit Seeding Labs Addgene Supports Rare Disease Research Listen to Our Interview ...

...receptors. Scientific Reports, 10(1). https://doi.org/10.1038/s41598-020-78827-9 Breen, C. J., Raverdeau, M...Muthusamy, A. K., Liang, Y., Brown, T. A., Lemon, W. C., Patel, R., Lu, R., Macklin, J. J., Keller, P. J....protein function and cellular components: It's important to assess how dye-conjugation affects protein ... interpretation of experimental results. It's important to consider how dyes can influence receptor conformation...efficacy of these ligands can be affected by the incorporation of fluorophores, as seen in studies involving...wavelengths for green, red, and far-red dyes is important for intracellular fluorescence live imaging ​​(...from cells under certain conditions, which is an important consideration for assay design (Breen et al., ...

..., larval zebrafish react by performing a distinct C-bend to escape possible danger (see figure). Larvae...: 19805086. PubMed Central PMCID: PMC2764931. Portugues, Ruben, et al. "Optogenetics in a transparent ...

...allow the digestion to take place for an hour at 37°C. This should be enough time to completely digest the...overhangs, insert and vector purification, and, most importantly, excessive time requirements. FastCloning is ...

...T-regs) into their microenvironment by producing C-C motif chemokine 22 (CCL22) and TGFβ. T-regs are suppressive...still many important questions in this exciting field of research. One of the most important: How will ...commute to and invade multiple organs, and most importantly escape the immune mechanisms of the host. The...against cancer. Sipuleucel-T by the Dendreon Corporation is currently the only FDA-approved DC therapy...therapies themselves can be made more effective. Some important factors researchers are beginning to focus on ... of cell types. This cell heterogeneity is an important element of successful tumor progression. A tumor...), and ability to metastasize. There have been reports of clinical trials going amiss and patient deaths...

... Kunjapur, Aditya M., Philipp Pfingstag, and Neil C. Thompson. "Gene synthesis allows biologists to source...is evolving so fast that it is a really nice opportunity to study it and see what happens when technological...

... new scientific observations. References 1. Rago C, Vogelstein B, Bunz F. Genetic knockouts and knockins...Vaimberg EW, Donovan KF, Smith I, Tothova Z, Wilen C, Orchard R, Virgin HW, Listgarten J, Root DE. Nat ...of the gene with multiple gRNAs, but the most important is that you could avoid potential off target artifacts...you could run another gel that highlights the important results (for example, if you needed to make a ... Despite the shifting landscape of research opportunities, it’s an exciting time to be in the lab. Based...

...protein of interest with a fluorescent reporter on the N- or C-terminus of the protein. One downside of...doi.org/10.1002/cpim.40 Picot J, Guerin CL, Le Van Kim C, Boulanger CM (2012) Flow cytometry: retrospective...a pressurized buffer called sheath fluid and transported through tubes or capillaries to a laser. As the...

...Autoregulation and Yeast-Mammalian Circuit Transfer C. Collins Lab Quorum Sensing Ellis Lab GeneGuard Endy... modification, including shuttle, integration, reporter, and tagging vectors. Metabolism Browse plasmids...

...biochemistry. There, I had exchanged contact with Amy C., Addgene’s intern coordinator shortly after hearing...capstone project report and presentation, during the final week at Addgene. This report shared my work ...Coming from an environment with limited opportunities and support, puts into perspective just how much one...hearing her talk about Addgene’s internship opportunity taking place next summer.  I applied to Addgene ...to differentiate which clone had successfully incorporated Syntaxin3 into its backbone. Once we had gotten...

...light. Previous iterations of FLiPs only used N- and C-terminal dronpa fusions. With these various designs...back in with the Addgene blog for future award opportunities. Xin Zhou: Fluorescent Light-inducible Proteins...particularly notes their ability to be precisely spatio-temporally controlled as a key advantage over other inducible...Oligosaccharides (HMOs). HMOs are thought to play an important role in nuturing the development of beneficial...consisting of sfGFP fused to a His-tagged ABC transporter known to bind to the HMO of interest. Fatima ...

...depositing can be grown using standard conditions (37 °C, LB, antibiotics, etc.) and stored as glycerol stocks...drop-down menu to indicate whether the tag is on the N or C terminus and whether it was present in the backbone...be entirely verified by sequencing. Upload any support files that will make it easier for scientists to...

...Antonini, M.-J., Rosenfeld, D., Park, J., Garwood, I. C., Yan, W., Niroui, F., Fink, Y., Chandrakasan, A.,...included in them for downstream control – this is important later! Next, additional components are added -...times more flexible than the brain fibers and importantly distortion of the fibers didn’t affect their ...temperature proximal to the implanted fibers. As important as its capabilities are the practical considerations...

... I as this enzyme recognizes the DNA sequence 5´-(C/T)CCTT-3' and digests double stranded DNA at this ...salt causes arcing during electroporation which would cause the electroporation to fail). When incubating...limit (lower transformation efficiencies have been reported with longer incubation); however, you may need...

...PubMed Central PMCID: PMC6070312. 2. Stark, Jessica C., et al. "BioBits™ Bright: A fluorescent synthetic...encodes it. That DNA sequence then needs to be incorporated into a host cell (through a process called transformation...FD-CF reactions affordable to many schools. Most importantly, experiments for FD-CF reactions can be fun and...

... system is capable of robust expression of high G+C content genes, as validated through monitoring the...plasmids use “Cas9-mediated adenosine transient reporter for editing enrichment”, or XMAS-TREE, to identify...

...Lindenbaum P, Martin-Fontaine M, Broucque F, Robin C, Hebben M, Merten O-W, Blouin V, François A, Redon...sequencing and analysis approach can be easily incorporated into standard QC regimens and will provide scientists...

...on designing and using RNA scaffolds: Delebecque, C. J. et al. Designing and using RNA scaffolds to assemble... source library of characterized aptamers and supporting publication: http://www.freebase.com/base/aptamer...

...arrays (e.g. for spacers A, B, and C, you could obtain arrays A, B, C, AB, AC, BC, and ABC). Nielsen Lab...expressed, the gRNAs cannot be released, adding temporal and/or spatial control to the system. pSQT1313...plasmids, which contain various promoters and reporters, and subsequently inserted into a Cas9-containing...-in-one CRISPR/Cas9 vector system.” Scientific Reports 4 (2014): 5400. PubMed PMID: 24954249. PubMed Central... Find plasmids from this paper at Addgene. 10. Port, Fillip, et al. “Optimized CRISPR/Cas tools for ...

...to produce and distribute virus. Addgene is a 501(c)3 nonprofit and our plasmid service was started based...milestone. Viral service supporters We would like to thank the companies who supported this project with funding...and be ready to provide extensive logistics support for export and shipping on dry ice. The Business and...seed money was used to build a completely self-supporting service so that Addgene is not dependent on grants... in accelerating the Viral Service. With their support we are able to provide faster access to a larger...number of viral preps. Visit our Sponsors and Supporters Page Thank you to all of our generous, sharing...Addgene repository. Without these scientists and the support of their technology transfer offices, Addgene would...

...this may be due to different lipid compositions of C. elegans membranes, which might beneficially influence...ApoB-NanoLuc reporter by inserting the NanoLuc luciferase downstream of the apoB gene. Using this reporter, the...Cell, 2019. PubMed PMID: 30639098. LipoGlo: A reporter for measuring ApoB bound lipoproteins Article ...cells. These changes were larger than previously reported for other organisms, and the Gottschalk group ...