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We narrowed to 303 results for: hal.2

Showing: 161 - 200 of 303 results

  1. Four Factors that Differentiate the Stem Cell Field

    Type
    Blog Post
    ...Interview with Stem Cell Researcher Darrell Kotton 2. Stem cell scientists care about patients As any grad... Yamanaka explained that it took 50 people nearly 2 years to create the first stem cell line that could...trials for ALS.   I have an uncle with Parkinson's, a half-brother who is a quadriplegic due to a spinal cord...

  2. Cancer, Inflammation and Immunity - Harnessing the Body’s Defenses to Fight Cancer

    Type
    Blog Post
    ...represent a favorable state for tumor progression (Ref 2). Future treatments may need to target this immunosuppressive...cell 30.1 (2016): 147-160. PubMed PMID: 27411589. 2. Todoric, Jelena, Laura Antonucci, and Michael Karin...Immunologists are coming together to tackle the challenge of curing cancer in cross-disciplinary projects...

  3. Will You Be My Mentor? Finding and Asking for Mentoring Support

    Type
    Blog Post
    ...article in the Addgene Blog Mentoring series will cover 2 of the 7 mentoring questions I set out to answer. ...101-Building Relationships for Success”. Choose 1-2 tactics to try. One of my favorites is making the ...current position Understand your interpersonal challenges if they know some of the people you work with...

  4. Using Video to Share Your Science: We Share Science

    Type
    Blog Post
    ...Information Science and Technology 60.4 (2009): 858-862. 2. Van Noorden, Richard. "Interdisciplinary research...Librarianship and Scholarly Communication 2.2 (2014): 2. Additional Resources on the Addgene Blog 6 Tools...solutions to complex challenges. As a cross-disciplinary researcher, the challenge of keeping up on studies... Sharing your research with the world can be challenging. After months, or years, of grueling effort to...biology or international relations.  Many of the challenges in science today are too complex for any single...making publishing their research all the more challenging even when it is of quality). On the site you ...

  5. A New Generation of Adenine Base Editors Improves Editing in Primary Human Cells

    Type
    Blog Post
    ...NGG-3′ (“N” can be any nucleotide base, followed by 2 guanines). This limits the use of ABEs as DNA sites... in the promoter region of gamma hemoglobin (HGB1/2) leads to continuous expression of fetal hemoglobin...T-to-G•C base changes (Figure 1), but it can be challenging to make these base changes, especially in primary...𝛽-globin), which cause sickle cell disease and 𝛽-thalassemia, can be mitigated with the expression of fetal...

  6. 5 Tips for Troubleshooting Viral Transductions

    Type
    Blog Post
    ... digest and compare it to some reliable controls. 2. Viral stocks may be sensitive to freeze thaw cycles...PMID: 24003179. PubMed Central PMCID: PMC3760253. 2. Rohwer, Forest. "Global phage diversity." Cell 113.2...tasks, both of which are factors in the technical challenges they can present. Many researchers find themselves...after transfecting cells, replace the medium with half the volume typically used (e.g., use 5 mL medium...

  7. Deep Dive: Fixing and Permeabilizing for Immunofluorescence

    Type
    Blog Post
    ... of permeabilization and membrane integrity (Fig. 2). For example, saponin interacts with cholesterol ... at too high a concentration.      Figure 2: Examples of different permeabilization strategies...samples in a given state or point in time. You want to halt degradative processes while maintaining the structure...fixation-artifacts-and-how-to-minimize-them/  Goldenthal KL, Hedman K, Chen JW, et al (1985) Postfixation...

  8. FlipGFP, a novel fluorescence protease reporter to study apoptosis

    Type
    Blog Post
    ...parts: (1) β-strands 1-9 (β1-9) and the one ɑ-helix, (2) β-strand 10 (β10), and (3) β-strand 11 (β11). Each...activating caspase-3, and detected fluorescence within 2 to 5 hours, a time consistent with previous methods...fluorescence imaging of living organisms can be challenging due to issues with tissue autofluorescence, cell...

  9. Advice for Starting a Biotech Company from a Grad Student turned Entrepreneur

    Type
    Blog Post
    ...go anywhere. The team should initially consist of 2-3 individuals that are aligned in vision and outcomes... the technology. This transition takes around 1 – 2 years. The product is the measure of success. Technology...his co-founders raised $6 million. The biggest challenge during this time was learning to communicate effectively...

  10. Your Top Requested Plasmid in 2015!

    Type
    Blog Post
    ...PubMed Central PMCID: PMC4089965. 2. Sanjana, Neville E., Ophir Shalem, and Feng Zhang. "Improved vectors...only been available for a little over a year and a half, a small sampling of the research it has already...another great year of sharing!   References 1. Shalem, Ophir, et al. "Genome-scale CRISPR-Cas9 knockout...

  11. Fluorescent Proteins 101: Fluorescent Protein Timers

    Type
    Blog Post
    ...linear region to solve for total expression time. 2. Timer functionality with a single gene Although it...290.5496 (2000): 1585-1588. PubMed PMID: 11090358. 2. Tsuboi, Takashi, et al. "Age-dependent preferential...understand how protein localization changes over time.Hallmarks of fluorescent protein timers When FP timers...noticed a few properties that have now become hallmarks of ideal FP timers: 1. Ratiometric determination...

  12. Virus Protocol - Generating Stable Cell Lines

    Type
    Protocol
    ...Workflow Timeline Day 0: Seed and transduce Cells Day 2–3 (am): Remove media, replace with fresh media containing...Biological Safety Cabinet 0.5–10 µL single channel pipette 2–20 µL single channel pipette 20–200 µL single channel... 200–1000 µL single channel pipette Ice bucket CO 2 incubator Pipet controller Hazardous waste container...cell death, the cell media should be changed every 2–3 days to maintain the dose of antibiotic, which may...confluent 10 cm dish can be expanded into two 75 cm 2 flasks, etc. Pro-Tip This selection method results...genome-wide libraries for CRISPR screening. Sanjana NE, Shalem O, Zhang F. Nature Methods. 2014 Aug;11(8):783-...

  13. Plasmids 101: Repressible Promoters

    Type
    Blog Post
    ...immediate early gene expression. Genes Dev. 1988 Jun;2(6):718-29. doi: 10.1101/gad.2.6.718. PMID: 2843425...Targeted Gene Expression in Drosophila.” Genetics 186(2) (2010): 735-55. PubMed PMID: 20697123. PubMed Central... in Saccharomyces cerevisiae.” FEMS Yeast Res. 12(2) (2012): 197-214. PubMed PMID: 22129153. Additional...solve the problem of broad QF expression-induced lethality in flies. Riabinina et al. also developed chimeric...

  14. Our Most Popular Blog Posts from 2019

    Type
    Blog Post
    ...fusion location, there are many choices to make. 2. Prime Editing: Adding Precision and Flexibility to...and represent the diversity of tools we share on behalf of scientists. 9. Plasmids 101: Positive and Negative...fluorescent proteins at Addgene. 7. Designing Your Chalk Talk for the Academic Job Interview by Erik Snapp...blogger Erik Snapp breaks down how to prepare a chalk talk for an academic job interview including defining... research vision, organizing your vision into a chalk talk, and tips for practicing.  6. The Fluorescent...

  15. CRISPR Library Amplification

    Type
    Protocol
    ... recover, set up overnight growth (Estimated time 2-3 hours) Transformation should be performed at the... day to ensure that growth times are limited. Day 2: Harvest cells and purify DNA (Estimated time 3-4 ... Tubes should contain a total of 5 mL (3 mL SOC + 2 mL transformed Endura from two separate transformations...has been absorbed by the agar. This usually takes 1-2 minutes. Critical Be careful not to rip or shred the...incubation if needed. Place 100 mL sterile LB at 4 ℃. Day 2 (morning) Before beginning, prechill at least four...pellet. The total weight of each pellet should be ~1-2 g. Pro-Tip Make sure to weigh the empty tube beforehand...Quality Control (QC) Pooled libraries can be challenging and expensive to ensure adequate quality, but...

  16. CRISPR Activators: A Comparison Between dCas9-VP64, SAM, SunTag, VPR, and More!

    Type
    Blog Post
    ...gene activation, with some genes experiencing around 2-fold activation levels. Although other methods have.... This method has been shown to activate genes at 2- to 3-fold higher than SAM, SunTag, and VPR (Zhou ...Ter-Ovanesyan D, Haque SJ, Cecchi RJ, Kowal EJK, Buchthal J, Housden BE, Perrimon N, Collins JJ, Church ...

  17. PCR Applications for Copy Number Variation Assays

    Type
    Blog Post
    ...mean you have one copy of the gene; an RQ value of 2 means you have two copies, and so on and so forth....quantitative PCR. Expert Rev Mol Diagn. 2005 Mar;5(2):209-19. doi: 10.1586/14737159.5.2.209. PMID: 15833050...available, ddPCR may be a better option for more challenging cases.  Price point If budget is a concern...

  18. PITChing MMEJ as an Alternative Route for Gene Editing

    Type
    Blog Post
    ... PITCh vector via In-Fusion or SLIC cloning. Step 2: Design an insertion locus-specific gRNA The gRNA ...Find the plasmids from this publication at Addgene. 2. Dalvai, Mathieu, et al. (2015). “A Scalable Genome-Editing...cell lines can be created in about a month and a half, without the need for complicated cloning of homology...

  19. Fluorescent Proteins 101: Green Fluorescent Protein (GFP)

    Type
    Blog Post
    ...from GFP and the mutation definitions found in Table 2 may not apply. Table 1: The specific mutations comprising..., V163A EBFP F64L, S65T, Y66H, Y145F   Table 2: Functional role of specific mutations in GFP derivatives...and when they come into close proximity, the two halves of EGFP undergo folding, maturation, and fluorescence...

  20. Finding Your Science Policy Path

    Type
    Blog Post
    ...teaching and mentoring others (see more in Figure 2). Starting early demonstrates not only your interest...application process so you can learn more! Figure 2: The many ways to prepare for a science policy career...That’s what makes it so wonderful. The most challenging part of jumping into this career path from academia... your hair out, your successes, failures, and challenges you’ve overcome. This information may help you...completely new field without issue. This may be more challenging if you are trying to apply for an outright job...

  21. Better Dyeing Through Chemistry & Small Molecule Fluorophores

    Type
    Blog Post
    ...expressed as a fusion with your favorite protein and (2) a synthetic fluorophore-contianing “ligand” that ...cell 58.4 (2015): 644-659. PubMed PMID: 26000849. 2. Grimm, Jonathan B., et al. "A general method to improve... include: Self-labeling tags (e.g., SNAP-tag, HaloTag, TMP-tag) – These widely used systems consist of...

  22. Tips for Screening with Yeast Two Hybrid Systems

    Type
    Blog Post
    ...retain their individual activities when separated (2), and that DBD and TAs from different systems could... Enzymol. 1983;100:293-308. PubMed PMID: 6312261. 2. Keegan, L., Gill, G., and Ptashne, M. Separation ...functional luciferase only when fusions of the two halves of the luciferase protein interact and enable intein...

  23. Deciphering the Mysteries of Behavior with Viral Vectors

    Type
    Blog Post
    ...of these neurons via the channel Channelrhodopsin 2 (ChR2) reduced glutamate transmission from the mPFC...deliver Cre-dependent expression of  Channelrhodopsin 2 (ChR2), a modified receptor protein that activates...stay tuned!  References Alpár, Alán, et al. "Hypothalamic CNTF volume transmission shapes cortical noradrenergic...

  24. Illuminating Choices: A Guide to Selecting Fluorescent Dyes and Ligands

    Type
    Blog Post
    ... ligands can elicit varying degrees of β-arrestin-2 recruitment to receptors, indicating their agonistic...small-molecule fluorescent labels. Nature Methods, 19(2), 149–158. https://doi.org/10.1038/s41592-021-01338...interchangeable labeling using the HaloTag® system, the new Janelia Fluor HaloTag Ligands expand the self-labeling...these self-labeling proteins are SNAP-tag® and HaloTag®. These tags consist of a genetically encoded enzyme...Hayashi-Takanaka et al., 2014). Other dyes may prove challenging to stain intracellular compartments in in vivo...2017). These dyes were adapted into ligands for HaloTag® and are currently offered at Promega. Due to the...

  25. Custom CRISPR Screens & the Green Listed Software

    Type
    Blog Post
    ...six different research constellations: Doench/Root (2), Zhang (3), Wu (4), Yusa (5), Wang/Lander/Sabatini...Bioinformatics 33.7 (2016): 1099-1100. PubMed PMID: 28414855. 2. Doench, John G., et al. "Optimized sgRNA design to...that code for surface proteins. One of the first challenges with a custom screen approach is to identify ...PMCID: PMC4744125. 3. Sanjana, Neville E., Ophir Shalem, and Feng Zhang. "Improved vectors and genome-wide...essentiality profiling reveals gene networks and synthetic lethal interactions with oncogenic Ras." Cell 168.5 (2017...

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

    Type
    Blog Post
    ...Almo, Institute for Protein Innovation  1:18:45 - 2:01:18 (end) - Panel Alex Tucker (Ginkgo Bioworks)...their curation of citable, linkable figures. Cause #2: Hidden “negative” data We all want to be able to ...Pamela Hines (Senior Editor at Science) Edward J. Hall (Professor of Philosophy at Harvard University) ...experiments they’re reading about weren’t changed halfway through in order to chase a positive result. Beyond...

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

    Type
    Blog Post
    ...), 1668. https://doi.org/10.1038/s41467-019-09656-2 More resources on the Addgene blog CRISPR101: Validating... cancer immunotherapy targets but are embryonic lethal, hindering studies post embryogenesis. Using C-... in a developing or developed immune system was lethal (LaFleur et al., 2024).   Figure 1: Schematic...

  28. Bricking Science: Portraying Scientific Reality Through LEGO

    Type
    Blog Post
    ...motion videos is very time consuming; producing a 1-2 minute video takes an entire week of our free time...cycle of one step. Challenges in the LEGO science video process One major challenge when we started was...will bring people together and make these daily challenges a little easier to overcome. We want to encourage...

  30. QC Sequencing Technologies at Addgene

    Type
    Blog Post
    ...short-read sequencing are highlighted in Figure 2. Figure 2: Overview of short-read sequencing technology... up the pace Our QC team constantly faces the challenge of keeping up with the large amount of QC analysis...

  31. Twenty Years of Sharing: Addgene's Viral Vector Service

    Type
    Blog Post
    ...’re delivering their payload to.   Figure 2: Numbers of AAV tools, by category, available from... substrates, light, or engineered ligands (Figure 2). Biosensors, which include calcium and voltage sensors...depending on their serotype, are particularly challenging. Their efficacy in any given system can be affected...

  33. CRISPR 101: Which Cas9 Do I Choose for My CRISPR Experiment?

    Type
    Blog Post
    ...sequence is unique compared to the rest of the genome.  2) The target is present immediately upstream of a Protospacer...utilized allowing for PAM flexibility. Fig. 2: Prime editor engaging target DNA. The reverse transcriptase...essential genes whose knockout would cause cell lethality. The Krüppel associated box (KRAB) is a naturally...

  34. Quick, Versatile Plant Transgenesis with GreenGate Plasmids

    Type
    Blog Post
    ...Transgenesis." PLoS ONE. 8(12): e83043 (20 December 2013). 2. Carola Engler et al. "A One Pot, One Step, Precision...practical terms, what GreenGate can do is “save you half a year,” in comparison to more tedious and time-...

  35. Let There Be LITE Plasmids

    Type
    Blog Post
    ...First, the light-responsive molecules of cryptochrome 2 are the chromophores flavin-adenine dinucleotide and...and CIB1 – which they borrowed from Arabidopsis thaliana. The results are light-inducible transcriptional...

  36. Advice on Career Paths and the Green Card Process for International Researchers and Entrepreneurs

    Type
    Blog Post
    ... both substantial merit and national importance; (2) The foreign national must be well positioned to advance...outstanding achievements in your specific academic field, (2) that you have three 3 years of research experience..., Casseus Law suggests a mix of experts including 2 dependents –people who have worked with you i.e. your...for companies to submit an application on their behalf.  If an individual obtains a cap subject H-1B (...

  37. Microbes: Look, Listen, and Tell

    Type
    Blog Post
    ...similar optical quality (magnification of 140X and 2 micron resolution). I have worked many hours in a ...University). Right there on the floor outside the lecture hall. It was wonderful to see the ad-hoc ‘sciencing’ ...

  38. Addgene @ Keystone: Thoughts on Precision Genome Engineering and Synbio

    Type
    Blog Post
    ...much has been done with CRISPR technology in just 2 years and excitement about how much more there is ...systems, modifications to the RNA components, and challenging target genomes, such as the malaria-causing Plasmodium...Addgene depositor, discussed some of the many challenges of creating biological circuits, and I can only...

  40. Antibodies 101: Beyond Surface Labeling

    Type
    Blog Post
    ...would label everything but the CD8+ T cells (Figure 2). Even better, we can use the same fluorophore for...gating on the negative population.     Figure 2: Dump gating labels all unwanted cell lineages with...capacity and speed of cells. (a) With each division half of the dye molecules are lost and the fluorescence...
Showing: 161 - 200 of 303 results