We narrowed to 1,017 results for: PLE

...maintain itself within host cells.  The plasmid has multiple ori that are functionally differentiated from ...proteins from bacterial hosts. In a plasmid with multiple oris, one may function in one type of host and... the others are functional in other hosts. For example, the pJD4 plasmid harbors distinct origins (ori1... load (Meyer 2009). Figure 1: Idealized examples of a narrow host range plasmid with only one ori... ori and a broad host range plasmid with multiple oris that vary in DnaA-box and iteron structure, host-independent...Acinetobacter calcoaceticus, Actinobacillus pleuropneumoniae, Actinomyces naeslundii, A. viscosus, Aerobacter... hosts in which proteins can be expressed. For example, scientists can incorporate interchangeable antibiotic...

...coming weeks, but, in this post, we will outline a simple method for selecting the guide RNA, validating ...CRISPR Design tool supplied by the Zhang lab. This simple website invites you to input the target region ...happy with your selection, ordering the sgRNA is simple and easy. Several companies supply gRNA sequences...you gRNA in vitro first. In vitro screening is a simple and fast method for validating your selected gRNA...). Testing the efficacy of your gRNAs is then a simple matter of transfecting HEK293T cells with your ...perfect primer sets to use to genotype your pups. A simple PCR with these primers will let you know if there...-step generation of mice carrying mutations in multiple genes by CRISPR/Cas-mediated genome engineering...

...put, the majority of cell line problems come from people sharing cells: the black market of cell biology...when it is important to check their identity. For example, perhaps a working cell bank has been made by a...stocks of cells. Working stocks should be frozen in multiple vials and thawed to produce a new batch once you...the cells you are working with because you have plenty of maintained freezer stocks. It is vital that ...monitor any changes. Your tracking system could be as simple as a piece of paper on the front of the incubator...passage number, cell line, and date, which you complete each time you passage the cells (as well as recording...are changing but yet more is needed to convince people of some of the basic tenets of responsible cell...

... for using lentivirus, infect and select, seems simple, in actuality, many find using lentivirus to be...al. outline a do it yourself PCR-based approach complete with freely available positive controls. Routinely...labs prefer to collect and pool the harvests from multiple time points while others opt for a single harvest...transfer and packaging plasmids. A benefit of the multiple harvest strategy is that each day you are supplying...destabilize the viral envelope. One drawback of the multiple harvest approach is that it tends to result in...buffers used, and other experimental parameters. For example, while freeze thaw is generally not recommended...that lentivirus produced at pH 6 are resistant to multiple freeze thaw cycles. Consequently, it is best to...

...proteins. lon protease: Strains where this is completely deleted (designated lon or Δlon) similary reduce...eukaryotic proteins Lemo21 (DE3)* Chloramphenicol (pLemo) Rhamnose-tunable T7 RNAP expression alleviates...replication fhuA2 [lon] ompT gal (λ DE3) [dcm] ∆hsdS/ pLemo (CamR) Expression of toxic, insoluble, or membrane...variety of sources, so fear not—there are a few simple troubleshooting measures that can help get you ...will not express in an IPTG induction strain for example, nor will a p15 plasmid be compatible with a pLys...cell-density they support; however, minimal media supplemented with M9 salts may be preferable if the protein... Proteins which contain hydrophobic regions or multiple disulfide bonds may aggregate and become insoluble...

...good guidelines to follow in order to ensure that people can quickly and easily identify what your plasmid... empty backbone name in your plasmid name. This simple piece of information can often convey many important...insert abbreviation to specify what species it is. Example: ‘h’ is for Human (homo sapiens), ‘m’ is for mouse...plasmid and their relative position to the insert. Example, if you have a Flag tag on the N-terminal of your...directly before or directly after the insert name.   Example: If the Glutamine at position 295 was mutated to...pBACKBONE-Flag-hGene-EGFP or pBACKBONE-Flag-hGene(wt)-EGFP These simple rules allow any scientist to know what is in a ...

...a world of possibilities in the engineering of complex gene networks. A comparable system for targeting... of RNA is highly desirable for extending the complexity of genetic circuits, allowing for tighter spatio-temporal... of interest. The new tool was developed and implemented by applying the Golden Gate cloning method to...March is fused to a translational repressor. In principle, however, designer PUF proteins can be fused to...in addition to their use in basic biology, for example, by targeting pathways underlying cancer or other...Assembly Kit out for themselves, she does have a couple of tips: First, she recommends using standard BsaI...

...Institute for Biological Studies) Multiple uses: From multiplexing to multiscaling NIR-shifted spectrum...spectrum is perfect for use in spectral multiplexing (Shcherbakova et al., 2018). NIR FPs can be combined ...Verkhusha, V.V. Near-Infrared Fluorescent Proteins: Multiplexing and Optogenetics across Scales. Trends Biotechnol...11, 239 (2020). Oliinyk, O.S., Shemetov, A.A., Pletnev, S., Shcherbakova, D.M. & Verkhusha, V.V. Smallest...cyanobacteriochrome as versatile tag for spectral multiplexing. Nat Commun 10, 279 (2019). Oliinyk, O.S. et...Huisman, T.M., Verkhusha, V.V. & Hodgson, L. Direct multiplex imaging and optogenetics of Rho GTPases enabled...

...locations using Cas9 and track the localization of the complex. Broccoli, the other green fluorescent aptamer,... RNA. Fluorescence is observed when these RNAs complex and the entire aptamer can hybridize and fold. ...uniqueness of the photostability of the Corn-DFHO complex. For dessert (last but not least), the only fruit-named...fruit-named aptamer, Mango, binds to multiple derivatives of thiazole orange (TO) and fluoresces in the...Other groups have used Mango to study RNA-protein complexes (Panchapakesan et al., 2017) and short non-coding...mango aptamer-fluorophore: a bright, high-affinity complex for RNA labeling and tracking." ACS chemical biology...transcription using a photostable RNA–fluorophore complex." Nature chemical biology 13.11 (2017): 1187. PubMed...

...resources on self-promotion (how hard it is for some people, especially women), visibility (how to get it, ...especially if introverted ), and networking (how to get people to remember you). What better way to accomplish...Interviews: The All-Important Job Talk. 1. Watch other people give great talks We are not talking about lab ...simplicity of their slides and how the slides complement the spoken message. Learn from their level of...cause and effect. Stories can and should be very simple. The goal is to get the listeners to be thinking...on a slide but not reading the explanation. The people that need the information will find it. Experienced...particular question. I like to ask about how many people use plasmids in their research and then see how...

...Brainbow multicolor labeling system - a perfect example of the continued utility of Cre-lox. Check out ...events specified by three loxP variants, but the principles remain the same. The first fluorescent protein...than 3-4 colors due to the tandem integration of multiple copies of the construct (about 8 in Brainbow-1.0...-1.0 mice.) The combinatorial effect of multiple, independent recombination events leads to a rainbow ...Brainbow-2 each contained a “default” state; for example, Brainbow-1.0 expresses RFP when the construct ...system, Sanes and Lichtman also developed a complementary Flpbow construct that is functionally similar...13):2835-46. doi: 10.1242/dev.094631. PubMed. Multiplex cell and lineage tracking with combinatorial labels...

...DePace’s yearly planning meetings revolve around two simple worksheets, each of which is filled out separately...summarized briefly below (and available of in the supplement of this paper), help each person organize his...including major project goals and effort needed to complete them. These documents dictate the subject matter...concrete plan in hand, trainees are motivated to complete their work rather than overwhelmed or confused...meetings may not look the same in every lab, but the principles behind this technique are beneficial for researchers...Extracurricular Activities for a Strong Science Career Path 7 Simple Strategies to Resolve Conflict With Difficult Supervisors...

... and a mashed potato bar (complete with all the fixings), as well as plenty of delicious desserts. We ... 10-year Anniversary Gala celebrated all of the people who have helped Addgene grow and spread our mission...playing all night was hand-picked (like our plasmid samples!) by Addgenies. If you were there, you might have...Addgene over the last 10 years as well as the many people that make up the Addgene family. It was great to... it to fruition. The concept, though relatively simple, changed the course of scientific research. What...

...sensors Combinations of classical genetic tools, for example Cre/loxP or Flp/FRT, with AAV vectors are common...vectors, and post-translational regulatory elements. Examples of these approaches are given in Tables 3-7. 3...specificity of cell targeting even further. For example, injecting AAVs at one location can result in expression...that received information from a region. In the example shown in Figure 1, injected retrograde AAVs at ...neuroscience is becoming more and more versatile. To complement the wealth of genetically encoded tools available...only reagents, but also method information. To complement these efforts, we’re beginning to share data ...

...Scientists are excited by somewhat unusual things. For example, I am not that interested in meeting movie stars...astronaut Jim Lovell I was speechless (the club of people who have been in space is pretty small). Therefore...this over 30 minutes, which was fun to watch. A simpler 1 protein - 1 RNA system Drs. Dounda and Charpentier...into one RNA to direct cleavage the result was a simpler 1 protein - 1 RNA system.  Dr. Charpentier advocated...breakthrough. In fact, the panelists agreed that, for example, the technology is already available today to cure... anemia in stem cells but this can’t be used in people until we learn more about safety and more about...

...that plagues other red GECIs, so the team could multiplex PinkyCaMP alongside green fluorescent and optogenetic...an inverted topology of FRCaMP (itself based on mApple, which means it likely has the blue-light photoswitching... exceptional brightness, photostability, and multiplexing capabilities. bioRxiv 2024.12.16.628673. https...Biomolecular condensates are membraneless liquid "droplets" that can concentrate and sequester specific macromolecules...extensive validation of the cell and gene therapy, so please check it out! Find AAV-CRISPRa plasmids here! ... the Qi lab's website crisprkit.org to request complete kits or find protocols and other resources. Find...  Multiple Authors...

...engineered ESCs), and chimeric adults mated to produce completely transgenic offspring. Although powerful, this...about 100 bps into the run, it got “dirty”, with multiple sequence traces overlapping each other! When the... finally, we have a technology, CRISPR, that is simple in concept, straightforward to use, and robust ...missing or added at the broken ends and, as such, cripple the gene. However, if repair material is provided...enrichment and selection offered by mouse ESCs. For example, we often screen only 15-25 mice when generating... Wu X, Jiang W, Marraffini LA, Zhang F (2013) Multiplex Genome Engineering Using CRISPR/Cas Systems. Science...-Step Generation of Mice Carrying Mutations in Multiple Genes by CRISPR/Cas-Mediated Genome Engineering...

... had to get used to the workspaces, as well as complete the lab safety and cybersecurity training before...project is a major project given to each intern to complete and present on.    Figure 1: Addgene's...weeks to work on cloning the Syntaxin3 gene in a pLenti vector backbone. During those five weeks, I worked...obtain eight clones. We ran digested and undigested samples of the clones on an electrophoresis gel in order...a 100% successful clone of the Syntaxin3 in the pLenti backbone! Once everything was confirmed, I worked...Toj-Villagran is a rising senior in high school who just completed their summer internship at Addgene. They plan ...

... or condition of interest using a simple keyword search. For example, searching the term "breast cancer...results section, but you had to dig through the supplemental files to see the actual data. Now you’re wondering...datasets, which are often obscured in large supplementary data files, BioGRID ORCS makes it possible for...sgRNA-level sequence read scores. The original supplemental data tables available with the publication are...broadest possible scope of screens to be curated, for example phenotype screens with read-outs other than cell...

...built, literally, 'brick-by-brick' to introduce people all around the world to the lives of researchers...wrong. Whether it be a bad fridge freezing your samples, or a dysregulated water bath boiling your experiments...Unfortunately, unless they’ve worked in a lab, most people don’t get to experience this very human side to... is to help make science mainstream and to show people the very human struggles in the laboratory routine...sharing and laughing at these struggles will bring people together and make these daily challenges a little...that we step on them with bare feet), to connect people with science. And so, “Bricking Science” was born...