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We narrowed to 328 results for: hal.1

Showing: 41 - 60 of 328 results

  1. Live and Let Dye: Self-Labeling Protein Tags

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    Blog Post
    ...intracellular targets. Nature Communications, 7(1), Article 1. https://doi.org/10.1038/ncomms10778 Erdmann...molecules in vivo. Nature Biotechnology, 21(1), Article 1. https://doi.org/10.1038/nbt765 Lavis, L. (2016... envelope design. Nature Communications, 14(1), Article 1. https://doi.org/10.1038/s41467-023-40788-8 ... capping protein. Nature Communications, 14(1), Article 1. https://doi.org/10.1038/s41467-023-39655-3 ...up linked to whatever molecule you choose (Figure 1). Just express the fusion protein and add a specific...proteins, opening up new applications.   Figure 1: Live-cell labeling strategies. The green structure...minimally-disruptive genetic tag on the protein of interest. Table 1: Summary of popular self-labeling tag systems ...

  2. Split Fluorescent Proteins for Studying Protein-Protein Interactions

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    Blog Post
    ...General Design Figure 1: Two proteins are fused to the FP(11) and FP(1-10) fragments. When Protein...and 11 to yield large (1-10) and small (11) fragments. In several cases an FP(1-10) fragment may be compatible...Description Article PI Blue EBFP2 EBFP2(1-10) and Capri(1-10) for use with GFP(11) Multiplexed labeling...Commun. 2017 Bo Huang mNeonGreen3 mNG3K(1-10) and mNG3A(1-10) for use with mNG2(11) Improved yellow-...sfCherry2 sfCherry2(1-10) and sfCherry2(11), also photo-activatable variant PAsfCherry2(1-10) Improved split...brings the pieces of the FP back together (Figure 1). This method is generally known as Bimolecular Fluorescence...even from a different base FP (for example, EBFP2(1-10) can be used with GFP(11)). Some of the articles...

  3. Antibody Validation for Flow Cytometry

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    Blog Post
    ...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...through their HLDA workshops. Let’s use TIM-1 (Single pass type-1 membrane protein), which has been designated...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...

  4. Lentiviral Vector Uses and Overview

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    Blog Post
    ...HIV-1 were transduced with a lentiviral vector containing an antisense sequence against the HIV-1 envelope...Trono and collaborators turned to the lentivirus HIV-1, well-known to infect non-dividing cells. The first...plasmids. The packaging plasmid carried a mutated HIV-1 provirus that couldn’t package itself due to a few...plasmid encoded the desired transgene flanked by HIV-1 long terminal repeats (LTRs) that facilitate viral... targeting and tracing cells in the brain (Figure 1). 2nd generation lentiviral packaging system The graphic...infectivity, is often used as a substitute for HIV-1 Env, which can only infect CD4+ cells. 3rd generation...lentivirus in the comments section below! References 1. In vivo gene delivery and stable transduction of ...

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

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    ...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...

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

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    ...in the immune system. Nature Immunology, 25(1), Article 1. https://doi.org/10.1038/s41590-023-01689-6 ...developed immune system was lethal (LaFleur et al., 2024).   Figure 1: Schematic showing the X-CHIME... 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...genes in the immune system. Nature Communications, 10(1), 1668. https://doi.org/10.1038/s41467-019-09656-2... cancer immunotherapy targets but are embryonic lethal, hindering studies post embryogenesis. Using C-...

  7. Tips for a 1st Time CRISPR User (by a 1st Time CRISPR User)

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    ...Pro-Tips: 1.Designing primers against genomic DNA to amplify the gRNA targets was challenging because you...The entire process from start to finish took about 1 month. Cas9 delivery Overview: Before any editing...lentiviral vector, lentiCas9-Blast (52962-LV, Figure 1). Some cells were then used to generate a monoclonal...rest of the cells were frozen down. Figure 1: Cas9 expression in A549 cells transduced with lentiCas9...Western blotting is a loading control. Pro-Tips: 1. It’s all about the MOI. In retrospect I used a way... pool had wildly variable Cas9 expression (Figure 1). This is a problem because Cas9 expression affects...cells were transduced (MOI = 37) and selected with 1 µg/mL blasticidin for 9 days. Single cells from a ...

  8. Harnessing TnpB for Genome Editing: A Compact and Versatile Tool for Your Research

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    ... Oligos Oligo 1: Add 'tcaa' to the 5′ end of your forward guide sequence (see Figure 1). Oligo 2: Add... Figure 1). Perform standard phosphorylation and annealing of the two oligos. Figure 1: Components... promoter for ωRNA (reRNA) expression (see Figure 1). Both plasmids contain a hygromycin resistance gene...provide a step-by-step guide for using a Dra2TnpB. Step 1: Design the 20-nt Guide Sequence Identify a TAM site...protocols (pp. 245–260). https://doi.org/10.1007/978-1-0716-4358-7_20  Karmakar, S., Panda, D., Panda, S... induction of meristems. Nature Biotechnology, 38(1), 84–89. https://doi.org/10.1038/s41587-019-0337-2...1166–1187. https://doi.org/10.1038/s41477-021-00991-1  Qiao, J., Zang, Y., Gao, Q., Liu, S., Zhang, X.,...

  9. Mouse Modeling, Part 2: Breeding and Crossing Mice

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    ...approximately 1 in 16 mice that harbor homozygous alleles at both of your genes of interest. About 1 in 4 mice...In Part 1 of our mouse modeling blog series, we covered techniques that can be used to introduce genetic...mice in your experiment. As you’ve learned in Part 1, there are many types of genetically engineered mice..., and assuming Mendalian patterns of inheritance: 1. Genotype your parental mice In order to ensure that...example of mice that are derived from crosses. In Part 1, we explained how genetic recombination in mice expressing...expressed, or inducible (this is explained more in Part 1 of this series). In any case, Cre is a dominant phenotype...that can impact reproduction include embryonic lethality, a short lifespan, or infertility and can arise...

  10. Degrading DNA with Cascade-Cas3

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    Blog Post
    ...Cascade's complexity. Figure 1: Schematic of Class 1 CRISPR types. Class 1 includes type I with a Cas3 ...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...this complex, developing Cas3 tools has been challenging. It can be difficult to ensure comparable expression...

  11. pCXLE toolkit: Efficient episomal plasmid-based method to reprogram peripheral blood cells to iPSCs

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    ...are based on the Epstein-Barr Nuclear Antigen-1 (oriP-EBNA-1) system where each vector contains a viral ...replication, OriP and the EBNA-1 sequence that encodes for the DNA binding protein EBNA-1 [7]. These two sequences...extra plasmid, pCXWB-EBNA-1 is used in the reprogramming cocktail. pCXWB-EBNA-1 lacks oriP and is unable...expressing additional EBNA-1 transiently. The transiently expressed EBNA-1 increases protein expression...pCXLE vectors listed below in Table 1 are available at Addgene. Table 1: List of pCXLE episomal plasmids,...obtained and the subsequent reprogramming success [1]. Fibroblasts need to be cultured and expanded in ...reprogramming factor cocktail, and viral protein EBNA-1. Methods for reprogramming hiPSCs from PBMCs While...

  12. Fluorescent Tagging of Endogenous Genes with SapTrap

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    ... 5’ and 3’ homology arm repair template (Fig. 1, Step 1). There is no need for PCR or cloning, as digestion...selectable marker, and connector to be released (Fig. 1, Step 2-3). Since donor plasmids of the same type ...correctly assembles the final targeting vector (Fig. 1, Step 4), and co-injection of the targeting vector...human cells. To use CRISPaint, users need 3 vectors:  1) a gRNA vector that targets the gene of interest, ... Tagging with Short Homology Donors.” Cell Rep.15(1) (2016):210-8. PubMed PMID: 27052166. Kunzelmann,...the tag for each insertion. Also, it remains challenging to screen for the genetically modified strains...fluorescent (EGFP, tagRFP, mCherry) and nonfluorescent (Halo, SNAP) tags, a selectable marker (floxed Cbr-unc...

  13. An Integrin Antibody Toolkit from IPI

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    ...-Integrin alpha 5 beta 1 [IPI-ITGA5/ITGB1.2] Integrin alpha-5,Integrin beta-1 Human, Mouse IgG1 Rabbit...-Integrin alpha 5 beta 1 [IPI-ITGA5/ITGB1.4] Integrin alpha-5,Integrin beta-1 Human, Mouse IgG1 Rabbit...specific integrins in cellular processes.   Figure 1: Integrins can shape into three conformations: (A)...immunoprecipitation and immunohistochemistry. Table 1: IPI antibodies  Antibody Target Proteins Reactivity...Human IgG1 Rabbit Anti-Integrin alpha M [IPI-LM2/1] Integrin alpha-M Human IgG1 Rabbit Anti-Integrin...Human IgG2a Mouse Anti-Integrin alpha M [IPI-LM2/1] Integrin alpha-M Human IgG2a Mouse Anti-Integrin... β1 integrin mechanics. Nature Communications, 13(1), 7471. https://doi.org/10.1038/s41467-022-35173-w...

  14. Negative Can Be Positive: Open AAV Data with Addgene

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    ...across scientific disciplines. Scientific Data, 8(1), 1–11. https://doi.org/10.1038/s41597-021-00981-0 Wright...excellent at infecting our target neurons (Figure 1). However, this one set of experiments didn’t provide...yet none of that data was published.    Fig. 1: Testing of three different AAV serotypes (AAV1, 8...purification and formulation. Molecular Therapy, 12(1), 171–178. https://doi.org/10.1016/j.ymthe.2005.02.021...Stimulation. Frontiers in Human Neuroscience, 14(December), 1–12. https://doi.org/10.3389/fnhum.2020.578687 Additional... peripheral nervous system presents a special challenge to scientists. What works well in one cell or ...vectors in neuroscience Let’s consider my first challenge when I started working with AAV vectors. I wanted...

  15. Treating Muscular Dystrophy with CRISPR Gene Editing

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    ...X-linked recessive disorder affecting approximately 1 in 5000 males. DMD is caused primarily by frameshift... applicable to 80% of DMD patients. Figure 1: Exon skipping strategies for Duchenne muscular dystrophy...needed. First, CRISPR delivery must be optimized to: 1. Reach a high percentage of muscle cells throughout...also be closer than we might expect.   References 1. Kaiser J. CRISPR helps heal mice with muscular dystrophy...neurogenetic diseases. Nucleic Acid Ther. 2014 Feb;24(1):69-86. PubMed PMID: 24506781. PubMed Central PMCID...);186-91. doi: 10.1038/nature14299. Epub 2015 Apr 1. PubMed PMID: 25830891. PubMed Central PMCID: PMC4393360...Precision genome editing faces the challenges above and more. One chief challenge is upregulating HDR in mature...

  16. SciComm with the Experts at Science in the News Part 1

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    ...Listen to the interview with Science in the News, Part 1 Podcast breakdown Introduction to Vini, Amy, and ...Communication The Purpose of Science Communication #1 Mistake Scientists Make when Communicating with Non-Scientists... This is the first half of a two-part interview with Vini Mani and Amy Gilson from Science in the News...eager to communicate their science. In this first half of our two-part interview with Vini and Amy, we’...

  17. Delivery Methods for Generating iPSCs

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    ...) element and the cis-acting EBNA-1 from Epstein-Barr virus. EBNA-1 binds oriP and allows for replication...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...Methods figures were adapted from this reference. 1. Malik, Nasir, and Mahendra S. Rao. "A Review of the...

  18. Quick Guide to Working with Drosophila Part 2: Controlling Gene Expression in Flies with Gal4/UAS

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    ...transcription activation system co-opted from yeast (1). It is a Drosophila geneticist’s main workhorse to...he still reads “#” as a “pound symbol”. References 1. A. H. Brand, N. Perrimon, Targeted gene expression... A Fly Geneticist’s Swiss Army Knife, Genesis 34, 1–15 (2002).PubMed PMID: 12324939. 3. S. E. McGuire,...Versatile Toolkit for Gene Expression in Drosophila, 3, 1–10 (2015). PubMed PMID: 21356876. 5. J. Chow, Z. Marka...two genes by one Gal4 driver will produce roughly half as much protein as overexpression of each gene individually...384–391 (2004). PubMed PMID: 15262411. 4. T. D. Southall, D. A. Elliott, A. H. Brand, The GAL4 System :...Marka, J. C. Kagan, Environmental Stress Causes Lethal Neuro-Trauma during Asymptomatic Viral Infections...

  19. Great Results Start with Great Standard Curves

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    ...    Figure 1: A 1 mg/mL control is diluted in a series ranging from 1:2 to 1:32. Each individual...example, Figure 1 depicts a 2-fold dilution series. Each step of the series dilutes by 1:2 for a series...series that ranges from 1:2–1:32. The specific dilution series used will depend on the expected concentration... points fit the trendline. R2 ranges from 0 to 1 with 1 being a perfect fit and 0 indicating that there...individual step of the series is consistent at 1:2. When preparing a dilution series, use a new pipette tip for...allergy testing lab, and the grad student down the hall from you have in common? All of them rely on standard...

  20. Scientific Reproducibility - Focusing on Solutions at the Minisymposium on Reproducibility

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    ... handed out at the event). Program 0 - 1:24 - Intro 1:24 - 29:31 - Reproducibility Overview - Jeffrey... 49:45 0 1:18:45 - Reagent Development - Steven C. Almo, Institute for Protein Innovation  1:18:45 - 2...reproducibility As Jeffrey Flier points out in his talk (1:24 - 29:31), there are a variety of ways of defining...enterprise towards greater reproducibility. Cause 1: Inability to repeat previously performed experiments...the Institute for Protein Innovation (IPI) (49:45 - 1:18:45), there are also initiatives to create more ...found. Consider the hypothetical situation in Figure 1 where many scientists are performing similar experiments...Pamela Hines (Senior Editor at Science) Edward J. Hall (Professor of Philosophy at Harvard University) ...
Showing: 41 - 60 of 328 results