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TypeBlog PostPublishedMay 5, 2020, 1:15 p.m....ultimately flattening the curve, it has become evident that widespread testing is crucial in controlling this ...PCR using Taq Polymerase and then transcribed to provide RNA substrates for Cas13. Cas13, upon finding complementarity...RPA, this technique uses Taq polymerase which is widely available and less expensive. Instead of using ..., field-deployable nucleobase detection and identification using FnCas9 (FELUDA) Sensitivity: – 110 femtomolar...FnCas9, this is also a good way to diagnose single nucleotide variations. Also, FnCas9 cleaves in a range of..., field-deployable nucleobase detection and identification using FnCas9. https://doi.org/10.1101/2020.04.07.028167....org Get the basics about CRISPR in our CRISPR guide Browse our entire COVID-19 plasmid collection ...
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Targeting HIV-1 with CRISPR: Shock and Kill or Cut it Out?
TypeBlog PostPublishedMay 10, 2016, 2:30 p.m....Over 25 million people worldwide are currently infected with the lentivirus HIV-1. Today, HIV-1 can be...the virus doesn’t completely disappear; it just hides in latently infected cells. To truly cure HIV-1,...removal of the HIV-1 genome from infected cells, an idea that became much more realistic with the advent ...Cas9/gRNA expression. A pooled analysis found no evidence of Cas9 cleavage at predicted off-target sites...into a therapy, these papers present tantalizing evidence that an HIV-1 cure may be within our reach. Similar... disease that infects over 250 million people worldwide. As we’ve previously seen in the CRISPR field,... Learn to Use CRISPR Pooled Libraries for Genome-Wide Screens Additional Resources onf Addgene.org ... -
Plasmids 101: Gibson Assembly and Other Long-Homology Based Cloning Methods
TypeBlog PostPublishedMarch 1, 2016, 3:30 p.m....different ways to clone DNA fragments which have provided appealing alternatives to restriction enzyme cloning..., I highly recommend watching this entertaining video created by our friends on the Cambridge 2010 iGEM...fragments to incorporate such regions of homology. This video gives a helpful demonstration of how to use Snapgene...the process works best with fragments over 200 nucleotides. This may be because the exonuclease could chew... through an entire fragment shorter than 200 nucleotides before the annealing and polymerization steps...methods (Kahl 2013). However, Gibson assembly isn’t ideal for synthetic biology standards, which rely up on...Type IIS restriction endonucleases) to cleave DNA outside of the recognition sequence. This requires that... -
Evolution of Brainbow: Using Cre-lox for Multicolor Labeling of Neurons
TypeBlog PostPublishedApril 24, 2015, 2:39 p.m....deletions are defined using 2 loxP variants (only identical loxP sites can mediate recombination.) The deletions...Optimizing the system: Brainbow-3 Although Brainbow provided neuroscientists with the vast array of colors ...variety of fluorescent proteins to find those with ideal characteristics (low aggregation, high photostability...produces a minimum of 8 colors. The use of AAV provides spatial and temporal control without the need ...Drosophila and zebrafish. Brainbow in Drosophila has aided in the mapping of neural circuits, such as connections... tool to mark and follow single cells due to the wide array and high stability of colors. Sanes and Lichtman...further refinements of the Brainbow technique will provide important insights into the complicated physical... -
Oh, The Places You Can Go: Careers in Science Communication - Curriculum Development
TypeBlog PostPublishedNov. 1, 2017, 2:04 p.m....open a heart to look at the inside. We do a lot of interactives and videos, as well, to supplement the...addition to testing those experiments. “We also do videos demonstrating an experiment or a technique to students...gel [for] electrophoresis or doing a dissection video. I did that in January. This involves going in front...the courses encompass what they want to teach.” Besides working with students and faculty, Caitlin also...would be to take any opportunity in the lab or outside the lab: write!” Caitlin did this by editing 15...on Facebook. “I think this helped me say with confidence when I interviewed, ‘I write science. I edit ... -
Choosing Your Perfect Empty Backbone
TypeBlog PostPublishedAug. 19, 2014, 3:39 p.m....various conditions. The first thing you'll need to decide when running your experiment, is which vector will...will discuss some of the features you may want to consider as you make your choice. The first and most important...selection marker or antibiotic resistance. Next, consider the expression level of your construct: would ... lentiviral or a retroviral backbone. This is considered to be a more robust expression system and has...are several types of yeast expression vectors. Consider your applications and experimental design. You....1 - TRC cloning vector). Check out our complete guide to empty backbones for additional details and guidance...Plasmids 101 blog series Addgene's Molecular Cloning Guide & Other Plasmid Protocols List of common epitope... -
10 Ways to Share Your Science!
TypeBlog PostPublishedMay 13, 2014, 8:34 p.m....their interest in science more generally to people outside of academia. Here are my recommendations on some...magazines or newspapers or other places online, and consider how those stories are being told. Twitter can ...science blogs written by practicing scientists. 3) Consider starting up your own twitter feed or talking about...Close the gap between science and society 6) Consider participating in a future SciFund Challenge. While...motivated more by the desire to help close the widening gap between science and society by connecting ... Santa Cruz, has been turning scientists into confident and connected science writers since 1981. That... both your plasmids and your research stories. Consider practicing communicating your research by guest... -
Advice for Choosing a Research Project
TypeBlog PostPublishedApril 17, 2014, 2:45 p.m....interests. We've outlined some important factors to consider and summarize helpful advice from the successful...Achieve Success in Academia". Important factors to consider when choosing a research project After joining...and goals. The following questions can serve as a guide when selecting a research project. What will the... are available in order to realize your project ideas. What mentors are available for guidance, intelligent...fungal biology.” Connie Cepko advises students to consider their goal of PhD training when selecting a research... good judgement with the current technology to identify a problem, asking what approach will I take and...touching a pipetteman. Take the time to generate new ideas and think about them critically before diving in... -
History of CRISPR Cas - A tale of survival and evolution
TypeBlog PostPublishedNov. 25, 2013, 10:12 p.m.... nucleases from re-identifying it (3). Surprisingly, few genes have been identified that neutralize CRISPR...Addgene’s CRISPR Guide or find CRISPR/Cas plasmids for your research. Do you have a cool idea or application... process the CRISPR loci to generate RNA, which guide a Cas nuclease to the specified exogenous genetic...engineering. The diversity of the CRISPR/Cas systems provides powerful lines of defense against invading phages...technology? Let us know. References Ishino Y, Hideo, S, Makino K, Mitsuko A, Nakata A. J Bacteriol. ... -
Hot Plasmids June 2018 - Reverse transcriptase, nanobody, and protein-DNA interaction tools
TypeBlog PostPublishedJune 27, 2018, 1:16 p.m....through our hot plasmids articles. These articles provide brief summaries of recent plasmid deposits and ...structural understanding thereof have the potential to provide improvements to existing technologies based on ...PubMed PMID: 29915061. A new method to identify proteins occupying a specific genomic locus: Caspex... as an affinity tag. This ‘Caspex’ construct is guided by a pre-defined sgRNA sequence to a specific genomic...preprint posted July 4, 2017 A new method to identify proteins occupying a specific genomic locus: C-BERST... and H2O2, APEX2 labels electron-rich amino acid side chains (e.g. Tyrosine) with biotin. These biotin-labeled...spectrometry allowing a researcher to specifically identify proteins that are enriched in the 20 nm radius... -
In Vivo Biotinylation of Bacterial Fusion Proteins
TypeBlog PostPublishedJan. 25, 2018, 2:09 p.m....kinetics of the target ligand. Another important consideration is whether the affinity of an experimental protein-ligand...Proteolytic removal of the fusion protein could provide a more relevant kinetic analysis, but the trick...immobilization through biotin-avidin binding could provide a true measurement of high affinity (i.e. sub-nanomolar.... MBP has a similar function as GST in that it provides a method to express a foreign protein in bacteria...biotinylate the lysine residue in the AviTag. As a side note, we found that adding a TEV linker dramatically...that we couldn’t find an all-in-one plasmid that provided an easy way to biotinylate a fusion protein in...biotinylation and purification strategy outlined here provides a basis to determine binding kinetics using a ... -
PiggyBac-ing Through the Genome Editing Field
TypeBlog PostPublishedMay 31, 2016, 3:30 p.m....transposon? The piggyBac™ transposon was originally identified in moths in 1983, but wasn’t successfully used... that have negative impacts on fitness. One consideration when making a transgenic cell line with the ...the piggyBac™ system, these challenges can be considerably reduced. The piggyBac™ transposon system can...donor-plasmid, you could use a single-stranded oligodeoxynucleotide (ssODN) as the donor [6]. The problem with... his specialized knowledge to help researchers identify the right technologies and strategies for their...sites is cytotoxic for proliferating cells and provides a simple tool for conditional cell ablation.,”...Zinc-finger nuclease-induced gene repair with oligodeoxynucleotides: wanted and unwanted target locus modifications... -
25 Work From Home Tips for Scientists
TypeBlog PostPublishedMarch 19, 2020, 1:15 p.m....yoga classes. Get outdoors. Taking a walk or a bike ride is a great activity for both exercise and clearing... times of stress due to #COVIDー19. Receive and provide support while being safe. @GHP_HarvardChan — Shekhar... (individual development plan) from Science to identify skills to work on for the next steps in your career...networking events. Download Addgene’s Science Career guide to learn more about career paths and things you ... Everyone. Plan out new experiments carefully. Consider submitting a registered report, where peer review...is based on the methods, not the results. Also consider using this framework to design your future experiments...Technique. Co-work virtually. With co-working, the idea is not necessarily to chat, but to mimic your usual... -
"Build Your Plasmid": The Game - Play to Teach
TypeBlog PostPublishedJuly 17, 2018, 12:23 p.m....brilliant ideas on how to solve many therapeutic and pre-clinical problems! Who knows, perhaps the ideas spurred...students play, you can observe, answer questions, and guide them to productive strategies while noting any gaps...their knowledge. Practically speaking the format I decided on was to have two teams of 4 people face each ...with them. My impression was that they had clearer ideas about gene therapy in the following lessons. And...freely share your projects with us! This exchange of ideas will help improve life science and biomedical education...molecular biology educational resources Find protocol videos to teach basic lab techniques Plasmid Land: The... -
Visualizing Genomic Loci with CRISPR-Sirius
TypeBlog PostPublishedApril 18, 2019, 12:53 p.m....without changing them. Thus, it has been widely adapted for a wide variety of applications: base editing,...fusions localized to specific loci based on the guide RNA sequence. A year later, they developed CRISPRainbow...RFP) to the genomic loci of interest. Pairs of identical hairpins within one gRNA will result in a blue... chromosome-specific loci in the genome and the guides are relatively unstable. Figure 1: CRISPRainbow...deliver CRISPR-Sirius into cells. They named this guide sgRNA-Sirius-8XMS2 which will recruit the HaloTag...Find plasmids for your research Read our CRISPR guide ... -
Summer SciComm Series: A PhD in Science Communication
TypeBlog PostPublishedAug. 23, 2022, 1:15 p.m.... took classes in biochemistry, microbiology, epidemiology, and bioanthropology. Studying human health ...doing in the lab and the interactions I was having outside of the “ivory tower.” I’d spend the afternoon lovingly...survived this long without downing any dihydrogen monoxide (H2O). Studying science communication as a PhD...quality of the study design or by their preconceived ideas about the controversial medical issues. Participants...beliefs even though all three texts were otherwise identical. Likewise, when participants read an abstract ...in a behaviour called the “biased evaluation of evidence,” in which people (conscious or unconsciously)..., looking for reasons to dismiss them. There’s evidence to show that people with greater levels of science... -
The MTA at Addgene
TypeBlog PostPublishedJuly 23, 2024, 2:17 p.m....’ll be confident in what to expect. No system, including MTAs, is perfect, but it does provide consistency...1995: the UBMTA. Because biological materials are widely varied with many properties, having well-understood...Instead, we facilitate the signing of MTAs by the Provider and Recipient (in Addgene parlance, the depositor...plasmids are distributed under the UBMTA, which is widely used across academia. It’s our process that differs...to note that it’s the depositing institute that decides the terms of distribution for their plasmids. ...Webpage, help center articles, and our MTA YouTube video for more information. Happy transferring! Many... -
Viral Vectors 101: Inverted Terminal Repeats
TypeBlog PostPublishedOct. 19, 2023, 1:15 p.m.... AAV (rAAV) vector (Figure 1). Everything else outside the ITRs gets left behind. Figure 1: Recombinant....com. ITR structure ITRs are symmetrical 145-nucleotide sequences that flank the ends of the single stranded...Figure 2). Additionally, the ITR contains a 4-nucleotide Rep binding element (RBE) that serves as a binding...sequences. GC tails are often located immediately outside of the ITRs in the plasmid backbone and do not ...since analysis of individual reads allows for identification of non-dominant ITR sequences. Restriction...Capsids More resource on Addgene.org Addgene's AVV guide AAV Viral Preps References Earley, L. F., Conatser...Alexander, I. E., & Logan, G. J. (2019). A User’s Guide to the Inverted Terminal Repeats of Adeno-Associated... -
What Good Citizenship Can Do for Reproducibility in Science
TypeBlog PostPublishedNov. 7, 2019, 2:18 p.m....This post was contributed by Deborah Sweet, Vice President of Editorial at Cell Press. Almost everyone who...some point. Usually it comes up when a researcher decides on a new project and begins by trying to reproduce... various different versions of checklists and guidelines to help authors understand what they are expected...NIH) now look for data management plans and for evidence of sharing for grant renewal—but there is clearly...hiring, funding, and promotion processes, I’m confident that everyone’s enthusiasm for actually making...Sweet from Cell Press. Deborah Sweet is the Vice President of Editorial at Cell Press. She trained originally...editorial work to focus on a broader role as Vice President. Her current responsibilities encompass overall... -
The Crispr Babies Saga Shows the Need for Action, Not More Delays
TypeBlog PostPublishedDec. 20, 2018, 1:15 p.m....research from bench to bedside is, in addition to thorough ethical considerations, in making sure that ... our CRISPR guide Find CRISPR plasmids by function Check out our genome engineering guide ...scientists and the public together to build consensus guidelines, but there was never a specific timeline for ...shuffling our feet on necessary actions to review and decide on the future of genome editing technology. We ...Harley and Lexi, and a clutter ofoutdoors-only spiders, all of whom are named Charlotte, all of whom are...