Skip to main content
Addgene

Enter a sequence to perform a BLAST-based search of the Addgene plasmid sequence database.

Information

In Category

We narrowed to 1,009 results for: PLE

Showing: 301 - 320 of 1009 results

  1. Plasmids 101: Biotinylation

    Type
    Blog Post
    ...avidin can be added to your sample. Any free biotin-binding sites on this complex can bind to your biotinylated... proteome that are biotinylated. In E.coli for example, the BPL, BirA, is responsible for biotinylating...plague these animals even if their diet was being supplemented with biotin. Thus something in the egg white...discovered in the 1960s, strepavidin. The Biotin-avidin complex can then be used and/or manipulated for a scientist...immunoprecipitation experiments. Biotinylated proteins or complexes can be easily purified using columns or beads ... added to the reaction for easy detection. The multiple biotin binding sites found on avidin are also ...This creates a large meshwork of biotin-avidin complexes bound to your protein of interest leading to amplification...

  2. March for Science

    Type
    Blog Post
    ...scientists gather information about reality informing people and their governments on what outcomes are actually...earlier analogy, no amount of shouting will make an apple fall up. As scientist and politician Rush Holt said...and public opinion do not seem to take note. For example, 91% of active research scientists (scientists ...consequences While frustration and anxiety have inspired people to rally for science, others are concerned about... is not going change depending on the number of people supporting it.  3) Linking scientists to one particular...march. Who is marching and why? Who? All sorts of people are choosing to march on April 22nd. As of April...for science education and literacy as I believe people that can see and understand data themselves can...

  3. Antibodies 101: Normalization and Loading Controls for Western Blots

    Type
    Blog Post
    ...abundance, consider the following example gel. Figure 5: An example gel (left) with bands corresponding...western blot requires a standard curve with a pure sample of your protein of interest (POI). This approach... a perfect world, protein concentration in your sample would directly correlate to protein abundance on...calculate things about earlier ones. In practice, multiple sources of error (red) influence these relationships...not perturbed by the experiment Unfortunately, multiple reports suggest that common housekeeping proteins...are highly expressed in cells. This means that at sample concentrations appropriate to detect a less-abundant...be measured in your assay. This is influenced by sample concentration as well as by antibody dilution —...

  4. Antibodies 101: Beyond Surface Labeling

    Type
    Blog Post
    ...lympho-/splenocytes, you might only be interested in one of the various cell populations; for example, CD8...These methods can include labeling markers with multiple proteins/antibodies, targeting markers found within...beyond standard surface labeling. Pro tip! Many people use 'labeling' and 'staining' interchangeably to...antibody can originate from a range of species; for example, rabbit or mouse. In a second step, a fluorophore-conjugated...Otherwise, your secondary antibody will bind to multiple primary antibodies, yielding a false positive ...FACS achieves greater purity of your sorted cell sample than MACS. An effective strategy to sort out your...the permeabilization solution can be the same). Examples of such targets include transcription factors,...

  5. How to Negotiate a Successful Lab Start Up Budget

    Type
    Blog Post
    ...equipment, reagents, animals, glassware, and even people. When developing a budget, I suggest that you focus...consumables (such as pipet tips, glassware, etc), get a simple list from your current P.I., lab manager, or administrator...need to tap for your future research team. For example, if you’re going to need a tech because you’ll ...regions where the host institute is located. For example, salaries in Kansas versus salaries in NYC will... that have the means to support your work. For example, if your work requires specialized equipment, try...need.  Prioritizing resources in your budget The simple practice of putting together a resource budget ...rather than focusing on long term investments. For example, purchasing “used” equipment can save you money...

  6. Summer SciComm Series: A PhD in Science Communication

    Type
    Blog Post
    ... shape what people know and believe about the world. It is vital to understand what people know – or, ...like?  One of my more recent studies looked at how people perceive and evaluate peer-reviewed sources of ...called the “biased evaluation of evidence,” in which people (conscious or unconsciously) pick apart sources... to dismiss them. There’s evidence to show that people with greater levels of science literacy are especially...hands-on science communication. Between New Zealand’s multiple Covid-19 lockdowns, I held mask-making workshops...the difference between airborne and aerosolised droplet transmission and how different fabrics and mask...illness. I later helped run workshops where I showed people how they can make reusable cloth menstrual pads...

  7. Synthesized by Ginkgo Bioworks, Shared by Addgene: SARS-CoV-2 Plasmids for Many Expression Systems

    Type
    Blog Post
    ...minipreps for small batches of samples. As soon as that process is complete, QC scientists prepare Sanger...mutation.  Once the samples arrive, each group of plates will have select samples prepared and screened...collection in your experiments and published your data, please fill out this form and we’ll add your citation ...available. Plasmids are continually added as they complete QC and you can find all available plasmids listed... high copy backbone with a T7 promoter driving multiple combinations of WT, codon optimized, untagged,...screened by NGS to spot check the samples, the rest will be put into a status we call “Archived.” This status...for all several thousand plasmids at once. Any samples that are requested by scientists, will immediately...

  8. Antibodies 101: Buffers, Storage, and Conjugates

    Type
    Blog Post
    ...you need more than one?   Multiplex assays Multiplex assays involve multiple antibodies, each with a different... the antibodies time-sensitive, using precious samples, or being compared to other data in a manner that...this post, the conjugates, or signaling molecules! Please keep in mind that this is just a brief introduction...conjugate that can be activated to emit light with a simple kit, and is readily available on any number of ...ain’t broke, don’t fix it. HRP is easy to find, simple to use, not sensitive to light in its inactive ...detected in the same assay. This is one of the most complex subjects in antibody-based protocols - but thankfully... this is an Antibody 101 post, so we'll keep it simple. If you follow the two basic guidelines below, ...

  9. Plasmids 101: Yeast Vectors

    Type
    Blog Post
    ...cerevisiae yes - This can complement ura4- S. pombe, but the complementation is weak. yes   LYS2 L-lysine...no S. cerevisiae yes - This can complement leu1- S. pombe, but multiple copies are required. yes   TRP1...vectors? Yeast are eukaryotes and thus contain complex internal cell structures similar to those of plants...mechanism of homologous recombination to facilitate simple gene replacement/mutation. Yeast expression plasmids...selection markers.  The yeast origin of replication (Please note: This first section primarily pertains to ... including a copy of a functional gene which complements the host’s auxotrophy. When grown on media NOT...drug selection methods feasible in yeast as a complement or alternative to using auxotrophic markers. ...

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

    Type
    Blog Post
    ...twice a week. I have done this with people I work with and people from outside my work for almost 20 years...they know some of the people you work with Give more informed feedback (for example, they may be able to... and way of expressing it in meetings? The more people you meet, the more potential mentors you will identify...tactics to try. One of my favorites is making the pledge to have lunch or coffee with someone I don’t know...tap them for advice. As a future alum, these are people you can “cold call” and ask for some of their time...friendships or work interactions. Watch for the people that are already teaching you and see if there ...have a broad range of mentors you will always have people you can turn to for help and advice. If you missed...

  11. Kiran Musunuru on the Newest TALEN Genome-Editing System

    Type
    Blog Post
    ...with the same genetic background and do an apples-to-apples comparison of all those mutations. If you ...compares to his CRISPR/Cas9 system. Addgene: For people unfamiliar with this new TALEN kit, can you describe...especially well-suited for use in stem cells. (See the complete protocol at StemBook). Addgene: How are you now...particular variants or mutations have been reported in people, we can use this kit to knock in those mutations...disease process. In the Cell Stem Cell paper, for example, we generated hepatocytes to examine the effect...much all investigators. Our kit makes it extremely simple to design TALENs. It only takes two to three days... find several mutations in the same gene or in multiple genes, all of which seem to contribute to a particular...

  12. Plasmids 101: Golden Gate Cloning

    Type
    Blog Post
    ... The destination vector contains sites with complementary overhangs that direct assembly of the final ...Unique 4 base overhangs can be used to assemble multiple fragments - up to 10 fragments are commonly assembled...fragment in an inverse orientation. This allows for multiple DNA components (promoters, genes, terminators,... reaction. For detailed Golden Gate protocols, complete with helpful tips and tricks, see The Sainsbury...editing which allowed for the ordered assembly of multiple DNA fragments to create TAL effector nucleases...plasmid, but it can also be adapted to express multiple gRNAs. Addgene has two Golden Gate-based gRNA ... 7 gRNAs into one destination vector, making multiplexing easy.  Disadvantages of Golden Gate cloning...

  13. Hot Plasmids and Viral Preps - March 2021

    Type
    Blog Post
    ... SCF E3 ubiquitin ligase complex. In the presence of auxin, this TIR1 complex binds to the degron sequence...elegant system that allows for the assembly of multiple DNA fragments efficiently and accurately in a ...a nucleus -> DNA compaction around histones -> Examples of histone-modifying enzymes and reader-effector...assembly (backbones available at Addgene) with examples of potential synthetic chromatin mediators and...system will allow investigators to rapidly create multiple synthetic chromatin-modifying proteins quickly...protein degradation and quantification of this depletion using imaging-based measurements (Ashley et al...of your protein interest would also result in depletion of the FP, which can be easily visualized.  The...

  14. CRISPR 101: Targeting Non-Coding RNAs with CRISPR/Cas9

    Type
    Blog Post
    ...sequence can completely disrupt expression of the transcribed ncRNA (Jiang et al. 2014).  Simple knock-out...possibly be made up of. The answer is… drum roll please… non-coding RNAs! You probably didn’t see that ... However, removing large lncRNA fragments with multiple gRNAs poses the risk of disrupting adjacent genes...transcription of the ncRNA (Hazan and Bester 2021). For example, inserting a polyadenylation signal downstream ...interactions between ncRNAs and adjacent genes are complex and not yet fully understood, so it is important...is not effective for your experiments, there are plenty of other CRISPR-based options that can target ncRNAs...variants may also be useful for targeting ncRNAs. For example, the Cas12a variant can cleave single-stranded ...

  15. New Viral Vectors - Summer 2024

    Type
    Blog Post
    ...optogenetic tool called PdCO.  This is an "optoGPCR" (coupled to Go, so it's an inhibitory opsin!) and due to...AAV5 Optogenetics Yizhar New viral service with multiple serotypes pAAV_EF1a-DIO-PdCO-mScarlet-ER-miniWPRE...AAV5 Optogenetics Yizhar New viral service with multiple serotypes pAAV_hSyn-PdCO-EGFP-WPRE AAV1, AAV5...AAV5 Optogenetics Yizhar New viral service with multiple serotypes pAAV-CaMKIIa(0.4)-eOPN3-mScarlet-WPRE...AAV5 Optogenetics Yizhar New viral service with multiple serotypes pAAV-CAG-FLEX-rc [Jaws-KGC-tdTomato-ER2...AAV1, AAV9 Biosensors Li New viral service with multiple serotypes pAAV-hSyn-GRAB_OTmut AAV9 Biosensors... AAV9, PHP.eB Controls Feng New viral service, multiple serotypes Looking for more viral vectors? ...

  16. Important Considerations When Using AAVs

    Type
    Blog Post
    ...Level-1). Other possible tools, lentiviruses for example, are not approved for use in vivo use at the University...Manitoba due to their Biosafety Level-2 or 2+. Please Note: Despite the fact that AAVs themselves are...you can see, AAVs of each serotype can transduce multiple tissues. So how do you choose the right serotype...hippocampus and cortex) within the mouse brain (4). An example from the cortex is outlined in the table below....thawing. Although more stable than other viruses, multiple freeze thaw cycles can still reduce AAV titer....is possible to use AAV constructs with self-complementing structure (scAAV) (7). As single stranded DNA...mapping of circuits In our lab, we use AAV-DREADD complexes to study neural function. Often the reporter proteins...

  17. Sequencing Options for CRISPR Genotyping

    Type
    Blog Post
    ...techniques differ in important ways, but can also complement one another by providing both broad and specific...Mismatch Cleavage Assay Not Provided 5% Inexpensive, simple Low-throughput, low sensitivity Sanger Sequencing...investigator can be relatively certain that their samples don’t have off-target mutations if they remain ...the clinic. NGS-based methods provide the most complete information profile regarding putative off-target...counts making them fast and viable options for multiplex sequencing in many laboratories. Nevertheless,...sensitive as targeted amplicon sequencing. For example, GUIDE-seq seems to have a minimum detection limit...DSBs, doesn't require high read counts, fast multiplexing Requires delivery of dsODN (potentially toxic...

  18. Plasmids 101: Protein Expression

    Type
    Blog Post
    ...relatively easily synthesized, must be produced using complex mixtures derived from cells or using live cells... expression of mammalian proteins that require multiple post-translation modifications for proper protein...systems Insect cells can also be used to produce complex eukaryotic proteins with the correct post-translational.... These systems enable high expression of very complex, glycosylated proteins that cannot be produced ...expression. Protein expression in bacteria is quite simple; DNA coding for your protein of interest is inserted...pET, pRSET, Gateway pDEST, and pBAD vectors for example. Protein expression from each of these vectors ... expression systems, including the majority of complex post-translational modifications. Extraction and...

  19. CrispyCrunch: High-throughput Design and Analysis of CRISPR+HDR Experiments

    Type
    Blog Post
    ... CRISPR samples. We invite you to jump in and try it out, or take a look at our live examples: experiment...adhere to the same principles listed here. It employs biological best practices. For example, it ranks guides...November, we began using it internally. Guiding principles behind CrispyCrunch While CrispyCrunch may evolve...you can depend on it to stick to a few guiding principles that we've followed since the beginning. It'...if desired. It works as a whole or in part. For example, you can analyze experiments designed outside CrispyCrunch...active guide. With these constraints in mind, we implemented the following algorithm for optimal mutation ...Francisco. Before starting his career in software, he completed a MSc in Psychology at McMaster University in ...

  20. Chromoproteins: Colorful Proteins For Molecular Biology Experiments

    Type
    Blog Post
    ...amajLime, cjBlue, meffBlue, aeBlue, amilCP, tsPurple, and gfasPurple) (Figure 2). Figure 2: Chromoprotein...expressed and gives the cell a magenta (amilCP) or purple (tsPurple) color. Though the toxin is integrated in ...). Eight years later, scientists isolated pink, purple, and blue chromoproteins isolated from reef-building...just by looking at the cells on the plate. For example, if your plasmid contains a gene encoding a blue... Figure 3: pTOX plasmids expressing amilCP and tsPurple genes. Image: Lazarus et al., 2019. One such... lab is the Waldor lab who used the amilCP and tsPurple chromoproteins to create negative selection markers... and Tse2 toxins, respectively, and encode the tsPurple chromoprotein.  Find the allelic exchange vectors...
Showing: 301 - 320 of 1009 results