We narrowed to 928 results for: Por

... 91bp on the PAM-proximal side, consistently supported higher frequencies (up to 60%!) of gene editing... and the targeting guide RNA into cells by electroporation. Results from our lab and others (Lin et al...Kim et al 2014) have shown that this technique supports extremely high frequencies of genome cutting and...nucleofection as (in our hands) this technique supports gene disruption or gene replacement frequencies...

...severely cold conditions. To understand their unique importance, let’s begin with a few basics.  Mesophiles vs...understanding and appreciating their biology. The importance of psychrophilic enzymes From an industrial perspective...decided to create a hybrid system where we could incorporate cold-tolerance inducing genes within a regular...scientist from Addgene. The talk highlighted the importance of reagent repositories and the myriad facilities...

... phage promoter pR, preventing expression of a reporter gene. When light is present, YFI is inactive, ...preventing repressor cI synthesis and allowing reporter gene transcription to take place. Addgene also...promoter is right for me? Here’s a list of some important variables to consider when choosing an inducible...’re expressing a gene that may be toxic, it’s important that your inducible promoter not be too leaky....

...binding ability of its nanobody partner, so it’s important to insert AsLOV2 at a spot in the nanobody that...light to control signaling provides greater spatiotemporal resolution than pharmacological or genetic ...activated Ras/Erk signaling by using a fluorescent reporter for Erk kinase translocation. These results demonstrate...control protein activity Read about RANbodies, reporter nanobody fusions Learn more about using the secondary...

...nuclease for your experiments, there are three important factors to consider: site accessibility, specificity...potential for off-target cleavage should be an important consideration during any experiment. Sensitivity...stringent PAM specificity. This can be especially important for base editing techniques, where Cas9 must be...many options! Comic by Maya Kostman. Another important consideration when choosing a Cas9 is balancing...

...most important resource that Addgene provides the scientific community? Katz: The most important resource...beyond. In your perspective, what do you think are important considerations for the future of open science?...immensely. Paratore: I have two children active in sports, so I enjoy watching their games.  If I’m not at...

...Nature has evolved ways to prevent degradation of important cellular RNA molecules with post translational...your gRNA. Pro tip: modifications are typically incorporated at the termini of the crRNA molecule (usually...Frontiers in Genome Editing). This is particularly important in experiments in which viability is being compromised...Ramel, MC., et al. Photoswitchable gRNAs for Spatiotemporally Controlled CRSIPR-Cas-based Genomic Regulation...

...about the process that was used to create this important collection.  Creating the Plasmids To begin constructing...antibody (R-mAb) expression plasmids, allowing for important reagents that could have been lost to continue...sequences available from each plasmid webpage. Importantly, the methods developed by the Trimmer Lab for...why choosing the right secondary is of primary importance. PLoS One. 2012;7(6):e38313. doi: 10.1371/journal.pone...

...permanently integrate into the host genome or temporarily express whatever it encodes. This ability to ...straightforward across applications, while electroporation of a human, for example, isn’t a reasonable...allows you to infect cells and deliver cargo, it’s important to understand their differences to choose the ...PMC413431. 2. Bukrinsky MI, et al. “Active nuclear import of human immunodeficiency virus type 1 preintegration...

...scientific conferences. Conferences provide the opportunity to meet people who are interested in the same...the conference. Large conferences have many opportunities but can be overwhelming as far as meeting new...the conference The conference may start in the airport. If you see someone traveling with a poster roll...friendly so stick with those. Don’t miss social opportunities, but don’t overdo it (some of my best studies...

...plants are the foundation of life on earth, and opportunities abound to build better fuels, feeds, foods, ...areas in plant biotechnology that are ripe with opportunity. We’ve decided to jump into one of those unexplored...out their mission of creating new forums and opportunities for open innovation in plant synthetic biology...build the plant synthetic biology community and to support more efficient engineering of a better tomorrow...

.... But how do you find these elusive teachers, supporters and advisors? I tried to start a more formal ...for their development plans. They all found the support of the group to be intrinsic to their success. ... continue to meet regularly, providing ongoing support, advice and accountability for development goals...mentoring relationship.  Celebrate your successes and support each other as you transition to the next stage ...

...people can’t find you on LinkedIn, you will lose opportunities and hiring managers will think it is odd. For... you don’t add a personal note, you waste an opportunity to start things out right. Also, many people ...requests without a personal message and they may report you as spam or “I Don’t Know This Person.” If that...had a chance to proof read. This might not be important for a good friend, but if you are making a professional...

...microbiome by using engineered genetic tools and reporter microbes. Let’s take a look at a few of these ... because in vitro conditions did not optimally support growth of diverse species. These results further...is a ribosome binding site (RBS) followed by a reporter gene. The hairpin blocks translation by sequestering... reveals the RBS and allows translation of the reporter gene resulting in GFP signal. With the addition...

...containing Cas9 and two different gRNAs, and electroporated them with an ssODN repair template in induced...in cases of disease prevention. However, it is important to consider that already-tested CRISPR alternatives...clinical applications could be considered. It is important to consider the question of broader applicability... large step forward in the CRISPR field, it’s important to remember that the study hasn’t achieved any...

...2017). However, Acr inhibition is spatially and temporally ubiquitous, making them a rather blunt instrument...versatility opens up the potential for spatial and temporal control in many genetic targeting and editing ...rapidly screen thousands of proteins for one that can report electrical activity via imaging. They took the ...response to light, offers a way to spatially and temporally control cellular processes in vivo. Many photoactive...

...spectra. DsRed is the permanent fluorescent tag portion of Rosella: it will emit red fluorescence regardless...mechanisms behind autophagy by tracking what’s being transported to the yeast vacuole (i.e. cytosol, mitochondria...2008). Rosella: A fluorescent pH-biosensor for reporting vacuolar turnover of cytosol and organelles in...using a single fluorescent biosensor. Scientific Reports,5(1). doi:10.1038/srep12397. PubMed PMID: 26215030...

...viral Rev gene from the packaging plasmid and incorporating it into a fourth plasmid. In addition, the promoter...third generation packaging systems, this system incorporates the Tet-off transcriptional activation system...number of harvests will all impact the titer. Importantly, optimized conditions for one transfer plasmid...producing lentivirus, it is time to titer. One important consideration when titering a lentivirus is the...

...length can and should be varied as it can have important implications for the fusion protein effectivity...upon these original tools that allows for both spatiotemporal protein degradation and quantification of this...et al., 2021.   The AID system relies on a transport inhibitor response 1 (TIR1) expressed under a ...a desired promoter with a specific spatiotemporal expression pattern and an auxin inducible degron (AID...

...publication.   3’UTRome The 3’UTRs of mRNA are important for post-transcriptional regulation in C. elegans...combination of these methods provides unique opportunities to develop computational methods to better understand...system allows scientists to both spatially and temporally knock-down proteins of interest.  Plasmids for...intersectional strategy using a split intein for refined spatiotemporal transgene control in Caenorhabditis elegans...

...in a protein.    Cells expressing an RFP reporter plasmid for recombinase activity were exposed ...interest. However, FACS is unable to spatially and temporally target single cells. Now, Francois St-Pierre’...way to isolate individual cells with unique spatiotemporal phenotypes from large heterogeneous cultures...introduction & resolution of two DSBs in a fluorescent reporter. Find the plasmids here Yongsub Kim’s lab engineered...

...that meant rereading many of the original papers reporting various prime editing tools. These papers are ...editing application. And optimizing its sequence is important, as prime editing efficiency is affected by the...nucleotides. For longer templates, it is even more important to test different lengths and sequences, as unintended... cellular surveillance. To limit unintended incorporation of the pegRNA scaffold sequence into the genome...

...directly to buffer without a re-wetting step. Important: do not dry your membrane with antibodies on it...the ones shared by Bio-Techne, ThermoFisher, or Millipore Sigma. We’ll share the basic composition of these...Alternatively, companies including ThermoFisher, Millipore Sigma, and Azure Biosystems offer pre-made stripping...your western blot several times — some researchers report up to ten cycles! Just remember that a bit of sample...

...difficult to genetically edit due to toxicity from electroporation or poor cellular entry with other transfection...PAGE editing with viral sgRNA targeting mCherry reporter (mChe), with co-incubation with assist peptide.... CAP-Mac also robustly expressed fluorescent reporters in green monkeys and cultured human neurons, with...slightly reduced efficacy compared to AA9-X1.1. Importantly, AAV1-X1 is not neutralized by pre-existing antibodies...

...this rarely happens. But if you ever have an opportunity to read one of these structure papers – while...function in the human body?” … and probably most importantly… "how do we know this?” Over the past fifteen... nuclease active site wasn’t even close to its reported cleavage site on the complementary DNA strand....model also displays the several Lys/Ala mutations reported by Ian Slaymaker and colleagues to reduce off-...

....com.   The Cascade complex Cascade is an important component of the CRISPR Cas3 system. Most Cascade... appears to be conserved. This DNA degradation portion can be bypassed by leaving out Cas3 — a tactic ...is pulled through the Cas3. This mechanism is important for Cas3’s biggest trick. Cas3 likes to bluff,... — Cas3 will definitively degrade the targeted portion of DNA using only a single gRNA. Cas3 appears to...

...sensors are better classified by their fluorescent reporter and the mechanism of their voltage sensing component...rhodopsin proteins were initially evolved for ion transport, not fluorescence, they are not nearly as bright...for penetrating deeper tissues depths (1 mm), important for certain applications such as in vivo tissue...rhodopsin Ace2N and mScarlet. Ace2N quenches a proportion of mScarlet's emission via FRET.  Image from ...

...in-person portion got canceled, taking advantage of virtual meetings can still be a great opportunity to introduce...course of the last few years: recognizing the importance and benefit of remote and video communication...staff have become a new way to allow you to get important information about the campus even if you can’t...

...have been around since 2000 when Terskikh et al. reported the production of dsRed E5. This timer predictably... your experimental setup less cumbersome. 3. Portability across multiple systems Terskikh et al. showed...fluorescent timers that change color from blue to red report on cellular trafficking." Nature chemical biology...Chronology of islet differentiation revealed by temporal cell labeling." Diabetes 58.8 (2009): 1863-1868...

...suffers from several limitations. Perhaps most importantly, most Cas9 orthologs are less well characterized...(1, 3). The gRNA can be further modified by incorporation of an RNA hairpin, such as the MS2 hairpin, ...with nuclease-competent Cas9. An RNA hairpin incorporated into the truncated gRNA recruits a transcriptional...target site. Benefits of truncated gRNAs Importantly, the use of truncated guides has largely been...