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We narrowed to 250 results for: hal.2

Showing: 41 - 60 of 250 results

  1. Hot Plasmids: Spring 2025

    Type
    Blog Post
    Published
    June 3, 2025, 1:15 p.m.
    ...cell-attached recordings in primary visual cortex (V1) layer 2/3 (L2/3). B) Fluorescence from axonal boutons of neurons... generation and stable cell line creation (Figure 2). Looking for more? Explore our empty and EGFP retroviral... Lentiviral backbone plasmids here! Figure 2: Addgene cloned EGFP into pAG Lenti CMV N-HA Puro ...Glypicans are proteoglycans composed of a protein core, 2–4 glycosaminoglycans (GAG), and are (typically) tethered...either the optogenetic activator channelrhodopsin-2 or fluorescent reporter dTomato under the control ...important and abundant neurotransmitters, but it is challenging to monitor because only a small number of glutamate...to specific cell types in the brain can be a challenging — but critical — factor for successful experiments...

  2. Antibodies 101: ChIP

    Type
    Blog Post
    Published
    March 10, 2022, 3:25 p.m.
    ...the challenge, and take a few rounds of adjustments, to get everything just right!  Fig. 2: Goldilocks...to (1) run a DNA gel to confirm fragment size and (2)  incubate with RNAse and Proteinase K to purify the...proteins of interest to any DNA they were bound to (2) fragmented the DNA (3) used antibodies to specifically...such as sequencing or qPCR. Yay!  Where’s the challenge in that?  So while this method is certainly long...

  3. Plasmids 101: Dimers and Multimers

    Type
    Blog Post
    Published
    Jan. 10, 2023, 2:15 p.m.
    ...identification.   Fig. 2: Gel image of Plasmid 61564: pBAMD1-2 (~4.7 kb) digested with SpeI and... at ~3.6 kb and ~1.1 kb (lane 3). Uncut DNA (lane 2) has a band at <3 kb, which corresponds to the supercoiled...formation of plasmid multimers. Cell. 1976 Dec;9(4 PT 2):707-16. doi: 10.1016/0092-8674(76)90134-3. PMID: ...dimers in Escherichia coli K-12. Plasmid. 1989 Mar;21(2):138-41. doi: 10.1016/0147-619x(89)90057-7. PMID: ...Plasmids and Chromosomes. Microbiol Spectr. 2014 Oct;2(5). doi: 10.1128/microbiolspec.PLAS-0025-2014. PMID...Behavior of Plasmid DNA. Microbiol Spectr. 2015 Apr;3(2):10.1128/microbiolspec.PLAS-0036-2014. doi: 10.1128... PMID: 32338220. Crozat E, Fournes F, Cornet F, Hallet B, Rousseau P. Resolution of Multimeric Forms of...

  4. Technologies Enabled by NanoLuc® Luciferase

    Type
    Blog Post
    Published
    Feb. 8, 2018, 12:17 p.m.
    ...the system outlined in Schmid-Burgk, J.L., et al (2). In this post, I’ll cover two great applications ...molecularly evolved from mNeptune2 through 33 mutations and 2 deletions. CyOFP1 has a higher quantum yield, brightness...luciferin 24 hours later demonstrated a more than 2-fold brighter signal from Antares as compared to Fluc...PMID: 22894855. PubMed Central PMCID: PMC3501149. 2. Schmid, J.L., et al. (2016) CRISPaint allows modular...information. Many of our technology platforms like HaloTag® Fusion Protein, codon-optimized Firefly luciferase...concentrations of biological molecules. FRET sensors face challenges of photobleaching, autofluorescence, and, in ...cyan-excitable donors, phototoxicity. Another challenge to using FRET sensors comes when employing optogenetic...

  5. Early Career Researcher Toolbox: Social Media for Scientists

    Type
    Blog Post
    Published
    Dec. 11, 2018, 12:46 p.m.
    ...scientists are using social media professionally, and 2) I needed to start using Twitter. Sharing Chemistry...but my consistent writing for the Addgene blog (~1-2 blog posts per month) and use of Twitter and LinkedIn...PhD 👩🏾‍🔬💜 (@ThePurplePage) February 13, 2014 2. "The Scientist's Top 10 Guide to Twitter" blog post...you social media can positively impact your career, 2) given you tips for using various social media platforms...professional development, and while it’s not challenging to use, it’s hard to know where to begin. To ...

  6. FLEx Technology and Optogenetics: Flipping the switch on gene expression with high spatial and temporal resolution

    Type
    Blog Post
    Published
    Oct. 17, 2019, 12:28 p.m.
    ...information. References Abdallah, Khaled, et al. "Adeno-associated virus 2/9 delivery of Cre recombinase ...upstream of an opsin gene (e.g., channelrhodopsin-2, ChR2) fused to a reporter gene (e.g., mCherry) to...the correct orientation to be transcribed (Figure 2) (Sharma and Zhu, 2014). How does FLEx enable strong...Deniz, et al. "A FLEX switch targets Channelrhodopsin-2 to multiple cell types for imaging and long-range ...bacteriorhodopsin; a comparative study of strains of Halobacterium halobium differing in pigmentation." Biochemical...Rhodopsin-like protein from the purple membrane of Halobacterium halobium." Nature new biology 233.39 (1971): 149...opsin genes, and include bacteriorhodopsins, halorhodopsins, and channelrhodopsins, the first two found...

  7. Fluorescent Proteins 101: Luciferases

    Type
    Blog Post
    Published
    July 22, 2025, 1:15 p.m.
    ...its abdomen to produce yellow-green light (Figure 2). Despite the title of this post, this reaction is...; no chemical reaction takes place. Figure 2: Simplified chemical reaction catalyzed by luciferase... has a detailed list of luciferase options. Table 2: Properties of luciferase enzymes Name Natural ...Renilla reniformis), click beetle (Pyrophorus plagiophthalamus), deep-sea copepod (Gaussia princeps), Japanese...None No Click Beetle Red (CBR)(Pyrophorus plagiophthalamus) Natural or codon-optimized 613 (luciferin...

  8. Educational Planarian Tools for Teaching Developmental Biology Techniques

    Type
    Blog Post
    Published
    May 24, 2018, 1:20 p.m.
    ...neoblast cells and the gene Prohormone convertase 2 (pc2) labels the central nervous system. Evaluating... characterized in an unrelated worm (C. elegans) [2]. Delivering dsRNA matching a specific mRNA sequence...223). Link: http://abt.ucpress.edu/content/79/3/208 2. Fire, S. Xu, M.K. Montgomery, S.A. Kostas, S.E. Driver...experiments are either not accessible or too challenging for amateur scientists. Planarians, wormy creatures...available planarians: Girardia sp., Dugesia dorotocephala, Phagocata morgani, and Phagocata gracillis....differ. For example, Girardia sp. and Dugesia dorotocephalacan can be fed dsRNA-expressing E. coli that are...

  9. Adeno Associated Virus (AAV) for Cell and Gene Therapy

    Type
    Blog Post
    Published
    Nov. 7, 2017, 1:59 p.m.
    ...AAVs in the clinic Thus far, AAV vectors AAV1, AAV-2, AAV1-AAV2 hybrids, AAV-6, AAV-7, AAV-8, AAV-9 and...neutralizing antibodies against some AAV types, AAV1, 2, 3 and 5 are thought to be as high as 70% among the... 10.24 (2003): 1999-2004. PubMed PMID: 14566358.  2. Hirsch, M. L., et al. "Self-complementary AAV mediates... cargo (DNA or RNA) to target cells has been challenging. A variety of vector delivery technologies have...treatment appears to be both safe and effective. Challenges for AAV gene therapy Although AAV-based gene ...promising, scientists are working to overcome key challenges to their efficacy:  Pre-existing immunity: Many...

  10. Viral Vectors 101: An Introduction to AAV

    Type
    Blog Post
    Published
    May 30, 2023, 1:15 p.m.
    ..., AAV2/8 has a type 2 ITR and a type 8 serotype. Almost all vectors contain type 2 ITRs, which comes from...differentiated, non-dividing cells.   Figure 2: DNA rescue and transduction of a conventional single-stranded...quantitative PCR-based method for the titration of AAV-2 vector stocks."Molecular Therapy 6.2 (2002): 272. ...strand synthesis. In contrast, a scAAV vector with half the size of the ssAAV genome has a mutation in the...

  11. An Introduction to Adenovirus

    Type
    Blog Post
    Published
    Oct. 3, 2023, 1:15 p.m.
    ...either SARS-CoV-2, Adenovirus type 41 (AdV-41), or Adenovirus-Associated Virus type 2 (AAV2) – or some... was either 1) a physiological effect of SARS-CoV-2 toward destabilizing one’s natural immune response...response to Adenovirus (and/or to its partner, AAV) or 2) a sociological effect of the COVID-19 lockdowns causing...… Chiu, C. Y. (2023). Adeno-associated virus type 2 in US children with acute severe hepatitis. Nature... like other viral vectors, though significant challenges remain (Wold & Toth, 2013).  While the pre-existing...

  12. Pairing CombiGEM and CRISPR for Combinatorial Genetic Screening

    Type
    Blog Post
    Published
    April 12, 2016, 2:30 p.m.
    ...CombiGEM lentiviral vector backbone pAWp12 (Figure 2). Cloning from the pAWp28 storage vector to the pAWp12... before beginning your experiments. Figure 2: Strategy for assembling barcoded combinatorial gRNA...PMID: 25114216. PubMed Central PMCID: PMC4151723. 2. Doench, John G., et al. "Optimized sgRNA design to...2015; Koike-Yusa et al., 2014; Ma et al., 2015; Shalem et al., 2014; Wang et al., 2014), knock down (Gilbert... 26190106. PubMed Central PMCID: PMC4559080. 8. Shalem, Ophir, et al. "Genome-scale CRISPR-Cas9 knockout...

  13. Fluorescent Tagging of Endogenous Genes with SapTrap

    Type
    Blog Post
    Published
    Oct. 12, 2017, 2:26 p.m.
    ...digestion of the destination vector with SapI yields 2 sites- the first site accepts the sgRNA target sequence...marker, and connector to be released (Fig. 1, Step 2-3). Since donor plasmids of the same type will produce... a gRNA vector that targets the gene of interest, 2) a plasmid to specify the reading frame of the insertion...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...small epitope tags (HA, Myc, Strep tag II, AviTag, HaloTag, SpyTag). The Foerstemann lab developed a polymerase...

  14. Hot Plasmids and Viral Preps - January 2021

    Type
    Blog Post
    Published
    Jan. 5, 2021, 2:15 p.m.
    ... Opto-Cre-Vvd is fast and can cut in as little as 2 hours.  Sheets, MB, et al., ACS Synth Biol. 2020....they investigated three Casɸ orthologs, Casɸ-1, Casɸ-2, and Casɸ-3. This system is notable for several reasons...a minimal PAM requirement. For instance, for Casɸ-2 the PAM sequence is 5’-TBN-3’ (where B is G, T, or...deposited by Leopold Parts. This library delivers 2 randomly paired guides per construct and allows a ... is only ~70kDa, with a molecular weight roughly half of Cas9 or Cas12a. This is in part due to the fact...

  15. Antibodies 101: Flow Cytometry

    Type
    Blog Post
    Published
    July 20, 2021, 1:15 p.m.
    ...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, ...then flow cytometry can help you overcome these challenges and make these types of experiments easier and...fluorophores Fluorescent antibodies get around the challenges of fluorescent protein fusions by binding to ...

  16. Evolution of Brainbow: Using Cre-lox for Multicolor Labeling of Neurons

    Type
    Blog Post
    Published
    April 24, 2015, 2:39 p.m.
    ...Brainbow mouse neural labeling system. Brainbow-1 and -2 have been further refined to create Brainbow 3.2. ...M-CFP expression; these deletions are defined using 2 loxP variants (only identical loxP sites can mediate... each fluorophore. Third, Brainbow-1 and Brainbow-2 each contained a “default” state; for example, Brainbow...processes not previously visible with Brainbow-1 and -2. The general structure of Brainbow-1.0 is retained...First, the imaging of Brainbow mouse tissue was challenging due to low fluorescence intensity, caused partially...expression patterns. Hampel S, Chung P, McKellar CE, Hall D, Looger LL, Simpson JH. Nature Methods. 2011 Mar...

  17. Viral Vectors 101: AAV Variables That Matter

    Type
    Blog Post
    Published
    June 1, 2023, 1:15 p.m.
    ...testing your AAV.  Figure 2: Comparison of AAV serotypes 1, 2, 5, 6, 8, and 9 in two different ...) your experimental conditions and measurements; (2) the cells you’re targeting; (3) how much expression... Tropism and Axonal Transport of AAV Serotypes 1, 2, 5, 6, 8 and 9 in the Mouse Brain. PLOS ONE, 8(9),... Comparison of four promoters. Gene Therapy, 28(1–2), 56–74. https://doi.org/10.1038/s41434-020-0169-1...analyses of adeno-associated viral vector serotypes 1, 2, 5, 8 and 9 in marmoset, mouse and macaque cerebral...Adeno-Associated Viral Vector-Mediated Gene Delivery to the Hypothalamus. Human Gene Therapy, 21(6), 673–682. https:/...

  18. Synthetic promoter AAVs for cell-type specific expression in retinal cells

    Type
    Blog Post
    Published
    March 12, 2020, 1:15 p.m.
    ...retinal cell-type specific genes from mice. Strategy #2 (ProB): Create promoters using phylogenetically conserved...would express Cre-mCherry and GFP.   Figure 2: Synthetic promoters can be paired for targeted expression...1345-1356. https://doi.org/10.1038/s41593-019-0431-2  Neff, E.P. An AAV library for retinal cell expression... at the Institute of Molecular and Clinical Ophthalmology Basel recently took a different approach to ...to test for cell type-specific expression of GFP. Half (113) expressed in retinal cells four weeks post-injection...

  19. Twenty Years of Addgene Sharing: CRISPR

    Type
    Blog Post
    Published
    April 18, 2024, 1:15 p.m.
    ... were depositing and requesting each year (Figure 2). We did a similar analysis with CRISPR pooled libraries...different categories (Figure 2C). Note that in Figure 2, the numbers on the X axis refer to distribution and...rather than a cumulative total.    Figure 2: Addgene CRISPR deposit and distribution by function...rapid adoption of new tools, illustrated in Figure 2, and the longevity of tools seen in Figure 3, indicates... the easy part; using them can be a bit more challenging. Addgene’s free educational resources help scientists...

  20. Truncated gRNAs for Regulating Gene Expression

    Type
    Blog Post
    Published
    Jan. 10, 2017, 3:37 p.m.
    ...the absence of genome editing (Fig. 2 and 3) (5, 3). Figure 2: Gene activation with nuclease-competent...shown to exhibit more limited activity than SpCas9 (2). In many cases this is due both to lower relative...PMID: 24076762. PubMed Central PMCID: PMC3844869. 2. Chari, Raj, et al. "Unraveling CRISPR-Cas9 genome...different perturbations. Consider, for instance, the challenge of attempting to simultaneously and selectively...
Showing: 41 - 60 of 250 results