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TypeBlog PostPublishedNov. 14, 2024, 2:15 p.m....it is important to carefully select the target locations of knock-in studies to avoid off-target effects...it comes to targeting precision and practical application. If the CRISPR/Cas9 system is not effective for...lncRNAs with high specificity, particularly in applications requiring smaller delivery systems (Zibitt et...
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Making CRISPR Plasmids Using Fragmid
TypeBlog PostPublishedAug. 29, 2024, 1:15 p.m....ampicillin resistance cassette and an origin of replication, which are not schematized. Sizes of fragments...five months. When they tested those clones (in duplicate), 93% had the correct banding pattern in the restriction...correct. Because they created the assemblies in duplicate, they were able to confirm that at least one plasmid... -
Plasmids 101: FLEx Vectors
TypeBlog PostPublishedApril 28, 2016, 2:30 p.m....sites directs the Cre recombinase to invert, translocate, or excise a DNA fragment. The availability of... the function of various point mutations and truncations in the absence of the endogenous gene. Spatio-temporal..., allowing for clearer characterization of the location, morphology, and circuit mapping of a targeted... -
How-to: 5 Steps to a Great Panel Discussion
TypeBlog PostPublishedOct. 22, 2014, 1:57 p.m.... about making sure the audience is interested, educated and talking as a result of the content. Here are...contribute. The moderator should identify the funny, provocative or interesting stories that will make for a fun...sure all your panelists have the correct date, location and parking information and remind them about ... -
Tips to Make the Most of a Scientific Conference
TypeBlog PostPublishedMay 23, 2019, 12:26 p.m....instances. If your poster material is nearing publication, consider uploading your work to a preprint server...time to meet. Some conferences may also have a dedicated app with a social media component, but if it doesn...other posts on scientific sharing, scientific communication, and scientific networking Learn how to set ... -
Adenoviral Delivery of CRISPR/Cas9 Aims to Expand Genome Editing to Primary Cells
TypeBlog PostPublishedDec. 10, 2020, 1 p.m....and quiescent, post-mitotic mammalian cells. Applications of CRISPR delivery by adenoviral vectors Since...Strategies for Tissue-Specific Targeting. In: Applications of viruses for cancer therapy. Elsevier, pp ...Holkers M, Maggio I, Henriques SFD, Janssen JM, Cathomen T, Gonçalves MAFV (2014) Adenoviral vector DNA... -
Transferable Skills: Negotiation
TypeBlog PostPublishedMarch 3, 2022, 2 p.m....fingers or Ethiopian?) to where you are going on vacation (New York City or Hawaii?). So how can you approach...been: A job offer conflicts with a pre-planned vacation I’m looking forward to. How can I negotiate my... I’m excited while still being able to take my vacation? I want to learn more about a different area of... -
Antibodies 101: Conventional vs Spectral Flow Cytometry
TypeBlog PostPublishedAug. 3, 2023, 1:15 p.m.... tumors, and moreso than other methods, the identification and isolation of rare cell types. In recent...complexity index ranges from 0-1, with a value of “0” indicating those markers are completely distinct from one...one another, and a value of “1” indicating those markers are identical and completely indistinguishable... -
The AAVantages of AAV in CRISPR Screens
TypeBlog PostPublishedFeb. 4, 2025, 2:15 p.m....This reduces the chance of integration at random locations that could cause deleterious effects. It is important...multi-plasmid system, where you have one AAV vector dedicated to expressing Cas9, while the other houses the...integration into CRISPR-induced DNA breaks. Nature Communications, 10(1). https://doi.org/10.1038/s41467-019-... -
Of Myc and Men
TypeBlog PostPublishedJan. 19, 2023, 2:15 p.m....used in way more applications. Fig. 1 Making a Protein Tag (Abridged) (1) Locate a promising monoclonal... by its antibody without additional chemical modification steps and when the epitope was within the tagged... -
AAV Vector Quality Control: Going the Extra Mile with NGS
TypeBlog PostPublishedSept. 12, 2017, 1:44 p.m....These impurities cannot be removed during the purification step because they are inside the virus itself...very low concentration. So how do we make sure to catch all of these potential mishaps before distributing...Guerin et al., 2020). In addition to genome identification, we developed custom open-access Python scripts... -
Cloning Mammalian Cells with the Agarose Method
TypeBlog PostPublishedSept. 7, 2017, 12:17 p.m....to rapidly punch out cell clones with sterile truncated pipet tips. Indeed, an experienced operator can...wiped down with 70% ethanol, and a box of sterile truncated pipet tips (USA Scientific #1011-8410) is also..., and making sure we always maintain a backup replicates of each clone in case of contamination. We do... -
When is a Monomer not a Monomer? The Top Three Ways Your Favorite Fluorescent Protein Oligomerizes in Cells
TypeBlog PostPublishedApril 19, 2016, 2:30 p.m....problem for fusion proteins. One of the major applications of FPs is to visualize the localization, dynamics...even if they were described as monomeric in a publication. The gold standard in assays used to determine...An experimental study of GFP-based FRET, with application to intrinsically unstructured proteins. Protein... -
Which Fluorescent Protein Should I Use?
TypeBlog PostPublishedMay 20, 2014, 2:06 p.m.... to be considered when choosing an FP: Unique categories of fluorescent proteins Photoactivatable / Photoconvertible... used are GECIs - genetically encoded calcium indicators (e.g. GCaMP). Others include: pHluorin & pHTomato.... al. (2010) “Fluorescent proteins and their applications in imaging living cells and tissues” Physiol... -
AAVs in Retinal Gene Therapy
TypeBlog PostPublishedJuly 31, 2018, 12:55 p.m.... time advances in human genetics led to the identification of several disease-causing genes, including... But it is the hard work and persistence of a dedicated team of scientists led by Dr. Jean Bennett (UPenn...-1250015778 7. Yu W and Wu Z., 2018. In vivo Applications of CRISPR-Based Genome Editing in the Retina... -
Plasmids 101: Secondary Nanobody Toolbox
TypeBlog PostPublishedFeb. 27, 2018, 2:04 p.m....in bacteria and extracted with common protein purification methods. Read on to learn more about nanobodies..., Alexa Fluor 568, or Alexa Fluor 647 before application to the sample. Staining with this multi-color...PubMed PMID: 29263082 Find plasmids from this publication at Addgene. 2. Bates, M., Böhning, M.I., Chug... -
Plasmids 101: Choosing an Antibiotic Resistance Gene
TypeBlog PostPublishedOct. 10, 2023, 1 p.m....common antibiotic resistance genes and which applications are best (or worst) for each antibiotic. The...is expensive (relative to amp), and for some applications, the benefit it offers is marginal. Fortunately...antibiotic resistance ultimately comes down to your applications and needs. The more niche and specific your ... -
New Optimized Genome-wide CRISPRko, CRISPRi, and CRISPRa Libraries
TypeBlog PostPublishedOct. 4, 2018, 12:44 p.m....is only effective when sgRNAs target a specific location on the gene. For CRISPRa, sgRNAs were targeted...Calabrese pooled library and lentiviral prep here! Applications for the Root and Doench CRISPR libraries Overall...et al. "A protein-tagging system for signal amplification in gene expression and fluorescence imaging.... -
The Importance of a Fun Workplace: Company Culture at Addgene
TypeBlog PostPublishedSept. 3, 2015, 2:30 p.m....be an obvious one, but don’t make things too complicated for yourself (especially if you are the sole ...sole event planner)! If you have a very intricate itinerary, there is a higher likelihood that something ...Delegate- it will save you time and sanity! 10. Communicate Make sure people know that there is something... -
Controlling for Off-target Effects with a New Genome-wide CRISPR Screen Design
TypeBlog PostPublishedSept. 13, 2018, 1:55 p.m.... potential control, and provide evidence that truncated gRNAs have improved specificity with only a small...toxicity in high-throughput screens." Nature communications 8 (2017): 15178. Pubmed PMID: 28474669. Pubmed...Improving CRISPR-Cas nuclease specificity using truncated guide RNAs." Nature biotechnology 32.3 (2014):...