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We narrowed to 704 results for: des.1

Showing: 101 - 120 of 704 results

  1. Revamp Your Lab Meetings With Creative Virtual Collaboration

    Type
    Blog Post
    ...interaction fades away the longer we are on it – so it is important to take breaks. After 1 hour (or another...those ideas and build from those.   Figure 1: Virtual lab meeting? No problem! You can still creatively...salmon_phd References Frassl MA, Hamilton DP, Denfeld BA, de Eyto E, Hampton SE, Keller PS, Sharma S, Lewis ASL...in scientific research and in any workspace. Re-design lab meetings with a focus on discussion and brainstorming...pivotal in writing publications/grants too. As described by several works (Wuchty et al., 2007, Vicens ...another designated time) of meeting, someone could raise their hands or signal the approach of a brain...prepares a coffee, or simply moves away from the desk to switch off a few minutes. The host could also...

  2. Delivery Methods for Generating iPSCs

    Type
    Blog Post
    ...) element and the cis-acting EBNA-1 from Epstein-Barr virus. EBNA-1 binds oriP and allows for replication... cell reprogramming factors and wait for cells to de-differentiate. However it’s difficult, especially...Difficult to purify reprogramming proteins. Table 1: Key Features of Different Methods for Generating ...-based expression requires serial transfection of 1 or 2 plasmids that express the reprogramming factors...  Replicating oriP/Epstein-Barr nuclear antigen-1 (EBNA1) based episomes: These plasmids carry the origin...method Most iPSC research falls into two categories: 1) studies focused on better understanding the mechanisms...iPSCs experiences in the comments below! References  1. Sano, Masayuki, Minoru Iijima, Manami Ohtaka, and...

  3. AAVs CREATed for Gene Delivery to the CNS and PNS

    Type
    Blog Post
    ...nervous system (CNS). AAVs are popular tools because: 1) their genomes are easy to manipulate, 2) they have... stained for a neuronal marker). Check out tables 1 and 2 below for a summary of the quantified transduction... AAV vectors. What can these AAV’s be used for?  1. Multi-color labeling of neurons Multi-color labeling... dendrites extend to. The keys to this method are 1) high levels of color diversity so that cells are ...AAV.PHP.eB 69 +/- 4% 55 +/- 7% 76 +/- 5% Table 1: Transduction efficiency of AAV-PHP.B versus AAV-PHP.eB...expression into a two part system (figure 4d):  Part 1: A cocktail of three AAV vectors expressing RGB proteins...neuronal transduction efficiency, as shown in tables 1 and 2, with a cell-type specific promoter or enhancer...

  4. Troubleshooting and Optimizing a Western Blot

    Type
    Blog Post
    ...from 1:100–1:1,000. If your antibodies come from ascites fluid, try a dilution range from 1:1,000–1:100,000...the antibody’s dynamic range. Specifically, the 1:2, 1:1, and No dilution samples all have approximately...recommended dilution and/or range (typically between 1:500 and 1:10,000 for primary antibodies). You’ll want to...recommended dilution range is usually between 1:5,000 and 1:200,000. Check the manufacturer’s recommendations...bands, you may want to consult Tables 1 and 2 in our Technical Design of a Western Blot post to make sure...Membranes look the same from both sides, so it’s quite easy to think lane 1 is‌ lane 8. Common ways to keep...reagent (usually an antibody).      Figure 1: A protein gradient. The density of the bands, which...

  5. Typing CRISPR Systems

    Type
    Blog Post
    ...   Class 1 versus class 2 There are two CRISPR classes: Class 1 and Class 2. Class 1 CRISPR systems...archaea, though Class 1 remains largely underutilized compared to Class 2. Class 1 Despite their incredible...optimal plasmids to harness the power of Class 1 systems. Class 1 CRISPR systems can be further broken down...effector was developed, called Cas7-11. Despite being in Class 1, Cas7-11 is a single protein effector ...types, 33 subtypes, and 17 variants. Figure 1: Overview of the CRISPR classification hierarchy. ...effector complex after binding to a target. Class 1 is the most abundant of the two, comprising about ...incredible abundance, Class 1 CRISPR systems are less popular in the lab. This can be attributed to the multi-...

  6. Antibodies 101: Normalization and Loading Controls for Western Blots

    Type
    Blog Post
    ...pipetting or irregular membrane transfer. Figure 1: A loading control is a type of positive control necessary...because the loading control band is weaker in condition 1 than condition 2, the normalized values for the two...to using loading controls accurately. Assumption #1: The loading control is not perturbed by the experiment...without your knowledge? If you can’t trust Assumption #1, any change in the normalized intensity of your POI...interest band is significantly lighter in “condition 1” than “condition 2.” By eye, the loading control appears... the protein abundance axis, both blue “condition 1” points (POI and loading control) are smaller than...assumptions discussed above into question. Assumption #1: The loading control is not perturbed by the experiment...

  7. The Effect of COVID-19 on Liu Lab Plasmid Requests From Addgene Data

    Type
    Blog Post
    ...was around 33% of that observed between Jan. 1 and Mar. 1, 2020. Figure 2: A simple approximation...paper. Green and red lines depict the sum of mean +1 and -1 std for each paper, respectively. The fitted...complication in the China data – there is a period of 1 month, from 1/22/20–2/23/20, where there were zero plasmid... from 5/22/14 to 5/19/20). Each paper has between 1–32 plasmids and a total of 2–2,590 requests. We received...correspond to publications of popular papers. Figure 1: Plasmid requests by date. Parentheses depict the ...China. However, it is evident that if we believe that 1/22/20–2/23/20 are the more accurate dates of lockdown...are no exception to this effect -- meticulously designed experiments in labs around the world were put ...

  8. Opto-Nanobodies: Using Light to Manipulate Cell Signaling and Protein Purification

    Type
    Blog Post
    ...loops 1 and 6 resulted in light controlled binding, but with opposite effects: insertion into loop 1 caused...experiments.  AsLOV2 Inertion Site mCherry #1 mCherry #2 GFP Loop 1 dark-induced dark-induced dark-induced...favorite protein (YFP). You have two options:  Option #1: Affinity tag purification You tag YFP and use an ...and one against GFP. Insertion of AsLOV2 into loop 1 always generated OptoNBs with dark-induced binding... induced dark-induced blue-light induced Table 1: Summary of the effect of AsLOV2 insertion site on...purification is now a reality thanks to OptoNBs. Recently designed by the Toettcher Lab, OptoNBs are photoswitchable... put AsLOV2 because here it was less likely to destroy the nanobody’s function than if inserted into the...

  9. Plasmids 101: Biotinylation

    Type
    Blog Post
    ...7.1 (2014): 1–7. PubMed PMID: 26224555. PubMed Central PMCID: PMC4884002. 10. Boer, E. de, Rodriguez, ...attach proteins or polypeptides to a solid substrate (Holden and Cremer, 2003). References 1. Chapman-Smith... of the biotin-avidin interaction   Figure 1: Biotin is enriched in egg yolk while its binding ...matrices.” Journal of Nanobiotechnology 13.1 (2015): 1–14. PubMed PMID: 25980573. PubMed Central PMCID: PMC4461904...Northern blotting Cell surface labelling Attaching polypeptides to a substrate Flow cytometry/fluorescence-activated...your downstream application. Biotin labelled oligonucleotides can be used in a multitude of applications...

  10. Plasmids 101: Dimers and Multimers

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    Blog Post
    ...indicates the presence of a multimer.   Fig. 1: The monomer of Plasmid 12259: pMD2.G (depicted on...plasmid sequencing services using long-read sequencing (1-25 kb), such as Oxford Nanopore sequencing or PacBio...10017-26. doi: 10.1074/jbc.M110.176636. Epub 2011 Feb 1. PMID: 21285356; PMCID: PMC3060453. Bazaral M, Helinski...multiple circular DNA forms of colicinogenic factor E-1 from Proteus mirabilis. Biochemistry. 1968 Oct;7(10...Strausbaugh LD, Gray P, Vartak NB, Brown M, Talbot D, Berg DE. pBR322-derived multicopy plasmids harboring large...Methods Mol Biol. 2009;582:11-25. doi: 10.1007/978-1-60761-340-4_2. PMID: 19763938. Maucksch C, Bohla A...confirmations like linear and nicked plasmids, as described in this article.   Long-read sequencing Addgene...

  11. MXS Chaining

    Type
    Blog Post
    ...assembly. Journal of biological engineering. 2010;4(1):1. Pubmed PMID: 20205762. PubMed Central PMCID: PMC2822740...with minimal spectral overlap between them (Table 1). In each construct, 3 copies of the fluorescent protein... enabled labeling of particular structures (Table 1). Each construct was flanked with a CMV promoter (...construct was then introduced to HeLa cells (Figure 1). The resulting HeLa cells show robust labeling, with...fluorophore at the respective subcellular structure (Figure 1). Here, MXS Chaining gave researchers the ability ... all four constructs into one 15 kb insert. Table 1: MXS Constructs for Labeling Cellular Structures ...maxima Tethering partner Subcellular localization 1 TagBFP 399nm/ 456nm histone 2B (H2B) Chromatin 2...

  12. HA Frankenbody, a New Imaging Tool to Visualize Single Molecules and Nascent Peptides

    Type
    Blog Post
    ...HA-tagged proteins in vivo. Nature Communications, 10(1). PubMed PMID: 31270320. PubMed Central PMCID: PMC6610143...epitopes (Ewert et al., 2004). This scFv hybrid provides the probe structure and stability from the scaffold...translated quickly by the ribosome. This feature provides HA frankenbody the ability to bind nascent HA ...

  13. X-CHIME: Context Dependent Germline Knockout in Immune Cells

    Type
    Blog Post
    ...in the immune system. Nature Immunology, 25(1), Article 1. https://doi.org/10.1038/s41590-023-01689-6 ... strains and lentiviral expression vectors (Table 1). The four systems Name Function Plasmid ... GFP (vex) Rosa26-FlpO-ERT2; H11-Cas9 Table 1: X-CHIME systems Proof of Concept These systems were...was lethal (LaFleur et al., 2024).   Figure 1: Schematic showing the X-CHIME approach. Figure from...genes in the immune system. Nature Communications, 10(1), 1668. https://doi.org/10.1038/s41467-019-09656-2...LaFleur, et al. 2024.   The X-CHIME deposit also includes pXPR_071, a plasmid that leads to higher titer...blog CRISPR101: Validating Your Genome Edit How to Design Your gRNA for CRISPR Genome Editing CRISPR Cheat...

  14. Deep Mutational Scanning with One Pot Saturation Mutagenesis

    Type
    Blog Post
    ... mutagenic primers to introduce all desired codon changes (step 1 in the image above). A second universal...Supplementary Note 1 in Wrenbeck et al for details on how nicking was optimized. How should I design my mutagenic...making some or all of the following modifications: 1) increased concentration of Taq polymerase, 2) increased...introduced, and scalability (see Supplementary Table 1 of Wrenbeck et al). It uses a uracil labeled wild-...strand is nicked by the BbvCI variant not used in step 1. Then this strand is degraded with ExoIII and ExoI... the first digest (step 1 in above diagram). Remember that the BbvCI enzymes nick the DNA strand that ...time no, but there are 2 instances where it does. 1) When the backbone vector has multiple BbvCI sites...

  15. Important Considerations When Using AAVs

    Type
    Blog Post
    ...vectors without specific facilities (Biosafety Level-1). Other possible tools, lentiviruses for example, ...suspected cases of insertional mutagenesis in humans (1) and AAVs are non-enveloped viruses, so they are resistant...Subscribe to Viral Vector Blog Posts from Addgene 1. Location, location, location! What do buying real...important factors related to location. Location (1): What tissue? Location (2): Where in the tissue/which...the right serotype? Keep asking questions!  Table 1: AAV Serotypes and Tropism Experiment  Detection...the vicinity of the injection. Examining serotypes 1, 8 and 9, not only do they undergo retrograde and ...types of research may be enabled by the use of AAVs? 1) Combined functional and anatomical mapping of circuits...

  16. Antibody Validation for Flow Cytometry

    Type
    Blog Post
    ...workshops. Let’s use TIM-1 (Single pass type-1 membrane protein), which has been designated CD365 in the most...dye, respectively. WT and KO cells were mixed in a 1:1 ratio and fixed in 4% PFA and permeabilized in 0.1%...antibodies were diluted to 1 µg/mL except for 14558, which was used at 0.35 µg/mL (1/100). * = monoclonal antibody...specific antibody:target combination of interest (Figure 1). It can therefore be helpful to try all three when...to label an intracellular target.   Figure 1: HCT 116 WT and SYT1 KO cells were labelled with a...cells were stained with the indicated Synaptotagmin-1 antibodies and corresponding Multi-rAb CoraLite® Plus...between the wild-type and the KO population (Figure 1), further optimization can be performed to both maximize...

  17. Predicting Adverse Reactions to Monoclonal Antibody Drugs

    Type
    Blog Post
    ...immunogenicity score (Mattei et.al., 2022).    Figure 1: Generation of immunogenicity scores by predictive...Terry, F. E., Roberts, B. J., Rosenberg, A. S., & De Groot, A. S. (2022). In silico Immunogenicity Assessment...antibodies for the treatment of diseases. J Biomed Sci 27, 1 (2020). https://doi.org/10.1186/s12929-019-0592-z  ...in vivo animal models. In these cases, they are designed to mimic the human lymphatic system or even the....org/10.3389/fddsv.2022.952326Joubert, M. K., Deshpande, M., Yang, J., Reynolds, H., Bryson, C., Fogg,...

  18. Save Time with Transient Plant Leaf Transformations

    Type
    Blog Post
    ... medicinal plant, the Catharanthus roseus (Figure 1). The ultimate goal of our research is to enhance ... which one you choose depends on your experiment (de Ruijter et al., 2003). For example, we did our initial...Humana Press, 2009. 41-50. PubMed PMID: 19183892. De Ruijter, N. C. A., et al. "Evaluation and comparison...activities in seed plants." Plant cell reports 9.1 (1990): 1-5. PubMed PMID: 24226366. Lee, Min Woo, and Yinong...and a vacuum is applied with a vacuum bell or desiccator. Successful infiltration can often be seen by...

  19. When is a Monomer not a Monomer? The Top Three Ways Your Favorite Fluorescent Protein Oligomerizes in Cells

    Type
    Blog Post
    ...these situations in cells are described below (See Figure 1).   Figure 1: Each problem with dimerizing... in nature are prone to dimerization (i.e. EGFP) [1, 2] or even form obligate tetramers (i.e. DsRed) [...dimerizing fluorescent proteins is illustrated. 1) Membrane proteins fused to FPs that dimerize can form dynamic...active state due to closure by FP dimerization.   1. Transmembrane fluorescent protein fusions Transmembrane...bright, non-physiological, and difficult to ignore [1, 2]. 2. Fusion to an obligate dimer or oligomer Several... For a more detailed discussion see [5]. Problems 1 and 2 depend on protein concentrations. An imperfect... distance runner, gardner, and cook.   References 1. Snapp, E.L., et al., Formation of stacked ER cisternae...

  20. Degrading DNA with Cascade-Cas3

    Type
    Blog Post
    ...complexity. Figure 1: Schematic of Class 1 CRISPR types. Class 1 includes type I with a Cas3 effector...collection! A “complex” system Cas3 is part of the Class 1 CRISPR family, which means it requires multiple Cas...the brunt of the mechanical work for editing. Class 1 CRISPR systems make up the majority of CRISPR-based...CRISPR-based immune systems in bacteria and archaea. Class 1 is further broken down into different types, with ...making up type I, the most abundant type in the Class 1 category. These type Is can then be further broken...editing in human cells. Nature Communications, 10(1). https://doi.org/10.1038/s41467-019-13226-x Cameron...Cas11. Type I-E Cascades contains all five of these Cas proteins, while type I-C Cascades do not contain...
Showing: 101 - 120 of 704 results