We narrowed to 530 results for: abo.2

... the two DNA strands and releasing topoisomerase (2).  These days commercially available TOPO kits provide...Taq polymerase using your favorite PCR protocol.  2. Set Up TOPO Cloning Reaction: Mix together the PCR...PubMed PMID: 1658796. PubMed Central PMCID: PMC52876. 2. Novel approach to molecular cloning and polynucleotide...in mind that Taq polymerase has an error rate of about 1 in 3,500 bases. Typically polymerases with proofreading...

...non-specific proteins, and then elute YFP.  Option #2: Opto-Nanobodies (OptoNBs) purification You skip adding...blue lights. Which option do you choose?  Option #2 may sound like fiction, but light-controlled protein...experiments.  AsLOV2 Inertion Site mCherry #1 mCherry #2 GFP Loop 1 dark-induced dark-induced dark-induced...Addgene blog Learn about using nanobodies to control protein activity Read about RANbodies, reporter ...reporter nanobody fusions Learn more about using the secondary nanobody toolbox for immunodetection Resources...Find nanobody expression plasmids here Learn more about optogenetics in Addgene's optogenetics guide ...

...various components of the immune system. Figure 2: In-vitro models for immunogenicity prediction of ...drug reaction. This system would somewhat act like a 2-step verification. All approaches have their own limitations...Future Enhancements. Frontiers in drug discovery, 2, 952326. https://doi.org/10.3389/fddsv.2022.952326Joubert...determining the T-cell epitopes that will likely bring about unwanted immunogenicity. The 3-D models can be applied...limitations. Using methods together can inform decisions about in vivo application and clinical testing. This would...to drugs: adverse reaction terminology. WHO Collaborating Centre for International Drug Monitoring, Uppsala...

...Carballosa, et al, (2015) eNeuro, March/April 2015, 2(2) e0001-15.2015. Pubmed. Resources: http://openspim.org...orthogonal-plane fluorescence optical sectioning (OPFOS) (2). Arne Voie, David Burns and Francis Spelman focused...in LSFM. In LSFM the laser light sheet, typically 2-6  microns, illuminates only one thin plane of the...protein (GFP) (6).   The Dodt and Frank Bradke laboratories later developed the 3DISCO (3-dimensional imaging...

...indicated they wanted to see more negative data (Figure 2.) At the same time, when scientists who support open...is not only housed, but encouraged.    Fig. 2: Results from a survey conducted by Addgene in 2021...detailed methods and image descriptions, a minimum n = 2, and well labeled high- and low-magnification images...gene therapy trials. Human Gene Therapy Methods, 26(2), 71–76. https://doi.org/10.1089/hgtb.2015.040 Initiative...asked why they don’t share, many express concern about sharing negative data they cannot explain, perhaps...

... analogs such as 2-fluoroadenine (2-FA), organisms with functional APT will convert 2-FA to cytotoxic ...selection of plants undergoing RNAi, and LIIbeta F 1-2 RNAi, which allows assembly of intron-spliced hpRNAs...nucleotides, resulting in death. In the presence of 2-FA, potent gene silencing of APT is required for survival... “transitivity,” calls into question any claims about specificity of a given RNAi technique. While transitivity...conceptually similar to the microRNA precursor mentioned above and is a popular RNAi trigger in the mammalian research...substantial ways: variable silencing efficiency and labor-intensive processes to isolate RNAi plants. With...resource on the Addgene blog Browse our blog posts about plant biology Check out our CRISPR blog posts Resources...

...promoter and terminator flanking the CDS, and a Level 2 multigene vector with two other independently regulated...the sequence you are trying to clone. Figure 2: MoClo assemblies depend on compatibility of four-...sequentially for your Level 0, Level 1, and Level 2 assemblies using the plasmid sequence files provided...after each transformation. This could be as few as 1–2 colonies if you’re an experienced MoClo user, but ...Robert Hurt and Mohamed Soufi. So you’ve learned about the Modular Cloning (MoClo) system and what it can...these would be interchangeable between different laboratories. Therefore, choosing acceptor vectors and parts...Created with BioRender.com. The Sainsbury Laboratory Golden Gate Cloning videos made by the Nicola...

... 60,000 cups of coffee per day if everyone drinks 2 cups per day – which would mean ~ 300,000 cups of ...working, and networking at SfN, remember there are only 2 days left for you to make the most of this great opportunity...learn about all the amazing research being done with plasmids. I heard an impressive talk about the use...pulls off such an inspiring, thought-provoking, collaborative event – all while keeping the party going! At...scientists we've spoken with are excited and passionate about making such tools open-access. Though it is time...

...FLP and Cre for additional control   Figure 2: Expression of a morphological marker (eYFP) and synaptic...Sternson. 2008. “A FLEX Switch Targets Channelrhodopsin-2 to Multiple Cell Types for Imaging and Long-Range ...PMID: 18614669. PubMed Central PMCID: PMC2593125. 2. Buchholz, F, P O Angrand, and A. Francis Stewart....The FLP Recombinase of the Saccharomyces Cerevisiae 2 Micron Plasmid Attaches Covalently to DNA via a Phosphotyrosyl... on the Addgene Blog Learn More about FLEX-vectors Learn More about Cre-Lox Check out Retrograde AAVs...expression have been developed. Read on to learn more about FLP-FRT, Cre-lox, and how combinations of FLP and...

... will repeatedly nose-poke at the active hole [1, 2].    Place preference/place aversion also models reinforcement...spend most of its times in the opposite chamber) [2]. Laser activation is usually controlled automatically...two different ways 1) your hypothesis is wrong or 2) there is some problem with the technical implementation...PMID: 21716290. Pubmed Central PMCID: PMC3775282. 2. McDevitt, Ross A., et al. "Serotonergic versus nonserotonergic...of the important considerations you should think about when developing optogenetics behavioral experiments...scientific research is that the field benefits from collaboration and the exchange of ideas. The rapid spread ...versatility and standardization of genetic tools in the laboratory mouse, most optogenetics is performed in this...

...drastically with lower titers. As you can see in figure 2, the expression from the 5x1011 titer was essentially... roughly 6-fold increase in expression [3, Figure 2]. Note how expression increases exponentially with...international 61.1 (2002): S9-S15. PubMed PMID: 11841606. 2. Schultz BR, Chamberlain JS. Recombinant Adeno-associated...diluting - if you end up concentrating your AAV above your desired titer and want to dilute it, err on...

... 1 2010 3 Nature  3.3% 10.5%   3.2  9.6 2 2010 167  Nature 3.3%  10.5%  3.2  534.1 3...Microbiology  1.6% 1.2%  0.75   125.3 In figure 2, you can see the full impact from using these weighted...plasmids deposited at Addgene still accrue on average 2 to 3 times as many citations as other papers in the... with plasmids deposited at Addgene – typically about four times as many as papers without plasmids deposited...

...Nature Cell Biology, 26(2), 294–304. https://doi.org/10.1038/s41556-023-01342-2 Koblan, L. W., Arbab, M...order to bind DNA, Cas9 requires (1) a guide RNA, (2) a complementary DNA sequence, and (3) a protospacer-adjacent...region of separate strands called an R-loop (Figure 2). This allows the guide RNA to base pair with the ...accessible on the outside of the enzyme. Figure 2: CRISPR R-loop. Created with BioRender.com. Base...), 35–40. https://doi.org/10.1038/s41587-020-0592-2 Additional Resources on the Addgene blog CRISPR 101...still isn’t over! Keep an eye out for our next post about base editing, where we’ll tackle transversion editors... 101: Cytosine and Adenine Base Editors Read about four base editor reporters CRISPR 101: Prime Editing...

...fluorogen activating protein - dL5** in this case), and 2) a fluorogen (the TAP - targeted and activated photosensitizer...cells present 96 hours post treatment (see figure 2). Induced cell death was confirmed by TUNEL staining...PMID: 26808669. PubMed Central PMCID: PMC4916159. 2. Jacobson, K., Rajfur, Z., Vitriol, E., & Hahn, K.... in culture and in vivo.  Read on to learn more about this killer illumination technique! What is a photosensitizer...within the near-infrared (NIR) spectrum (>620 nm). Above is a brief schematic of how the MG-2I-dL5** FAP-...use MG-2I-dL5** with your research? Let us know about your experiences using photosensitizers in the comments... to Receive the Fluorescent Protein eBook Learn about Small Molecule Fluorophores Resources on Addgene.org...

...2023 (and at closer to 2 million plasmids shared than not!), we think it’s about time we updated this post...time does it take to make a plasmid? Figure 2: Survey results from a Twitter poll run in 2023, n...go wrong, replace them for free. Plasmids ship out 2-3 days after ordering, making it a toss-up as to whether...cloning before you can start your experiment? What about all the reagents you need to order? Sometimes, it...dive deep into generating the data you’re excited about. When we were closing in on the one million plasmids...choose the right cloning method Find blog posts about scientific sharing here Download the Plasmids 101...

...one of lox2272 sites will now be sandwiched between 2 loxp sites and is excised by Cre (Figure 1). This ...flank the transgenes (non-homologous pairs) (Figure 2.). They found that almost all off-target expression...exercise caution when using them.    Figure 2. Schematic diagram showing the mechanism of spontaneous... Tropism and Axonal Transport of AAV Serotypes 1, 2, 5, 6, 8 and 9 in the Mouse Brain. PLoS ONE 8:e76310... Pseudotyped with Viral Capsids from Serotypes 1, 2, and 5 Display Differential Efficiency and Cell Tropism...the most common concerns we get here at Addgene about this system is off-target or spurious expression...cells that don’t express Cre or Flp, what can we do about it?  Test the serotype and promoter The first step...

...lab found that H2B-GFP expressing cells exhibited a 2-4 fold increase in the number of endogenous indel ...Denham lab made several other small changes creating 2 other versions but these did not surpass GEmCherry2...fluorescence and in-frame fluorescence.  Figure 2: The GEmCherry reporter. Image from Højland Knudsen...off target effects. However, these methods can be labor intensive and quite time consuming.   As Cas9/gRNA...have been several systems to do this but most are labor-intensive, require multiple fluorescent proteins... Browse all of our CRISPR blog posts Find more about fluorescent proteins Get a refresher on CRISPR with...

...immunofluorescent images like in this paper (2). Metabolic assays such as Seahorse can be employed to look...are many docking sites available for integration. 2. How soon would you like your flies ready for use?...PMID: 16547094. PubMed Central PMCID: PMC1526508. 2. H. Jiang,  et al. Cytokine/Jak/Stat signaling mediates...Additional Resources on the Addgene Blog Learn about Using Cre in Drosophila Read Other Genome Engineering...

... working with is correct. It's also best to use 1-2 ug of the vector for digestion. PCR Design primers...plasmid; a starting point could be a molar ratio of 1:2 (plasmid:insert), but this parameter has to be optimized...colonies (5-10) and grow them in a small culture volume (2 or 3 ml) containing the corresponding antibiotic, ...they get old, they lose competency). If all the above is correct, you should probably have a second and...blog posts Read blog posts on plasmid cloning Read about different plasmid elements Resources on Addgene.org...

...termed fluorescence activated cell sorting or FACS (2). By analyzing single cells instead of the population...expression. A routine flow cytometry experiment uses 2 or 3 different fluorescent colors each measuring a...distinct populations on a FSC versus SSC plot (Figure 2).  A user can then “gate” around the specific population...cell and T cell into distinct populations (Figure 2). An immunologist who studies B cells, might then ... B cells, plasma cells or others.   Figure 2: Due to differences in their size and complexity, ...biosafety cabinet with cloning rings or following labor-intensive limiting dilution protocols? If this sounds...fluorescent microscopy must be used. To learn more about the most suitable fluorescent microscopy technique...