...insight to the most requested plasmid, the number of requested plasmids, or trends of requests over time...website, allowing depositors to download their plasmid request data, as a CSV file, anytime. The CSV is... Visualizing the data So, after downloading my plasmid request data, I started to explore how I could ...file is open for anyone to use.  View the Addgene Plasmid Request Data Dashboard! Figure 1: The ...

...cost required to make a plasmid. In 2023 (and at closer to 2 million plasmids shared than not!), we think...great Plasmid Debate of ’18, Blue Flame depositor John Doench calculated the time to make a plasmid. Doench... make a plasmid? Doench also calculated the cost of consumable reagents used to make a plasmid, which ... to make plasmids that already exist, and we significantly reduce costs of producing plasmids with high-throughput...lot of plasmids, it’s almost always faster to order from us!) Ask the PI If you want a plasmid for free...high-throughput” Instead of making one plasmid at a time, try constructing several plasmids in parallel. That way, you...collection pages to more easily find plasmids Search for a plasmid in Addgene’s repository ...

... a gene (cDNA) can be done by either plasmid or virus. A plasmid carrying your cDNA can be targeted for...resources on the Addgene blog Plasmid 101: Inducible Promoters Plasmid 101: Repressible Promoters CRISPR...knock out. Looking for some permanent knock out plasmids? Check out our Cre vector and Murine LoxP vector...of protein expression. Addgene has a dCas9 KRAB plasmid for transient knockouts!  Turn it on Turn it on...

...protocols, and, of course, plasmids. Today we’re trying a new way to present plasmid info with a new segment...The Hot Plasmids Segment. Click on the player below to listen to a quick (~5 min) Hot Plasmids podcast... new plasmid technologies from one of our recent newsletters. Listen to the entire Hot Plasmids episode...greatest plasmids through this podcast segment. Below, we break down each of the Hot Plasmids segments...articles on the Hot Plasmid webpage. We hope you enjoy this new way to learn about plasmid technologies. If...comments about the Hot Plasmids Podcast Segment, or have recommendations for new Hot Plasmids topics, let us ... comments section below. Hot plasmids covered in this segment Hot Plasmid 1: CRISPR X from Michael Bassik's...

...most request plasmid in 2016 was... pSpCas9(BB)-2A-Puro (PX459) V2.0       Similar to last year, the top...for sure, as is obvious from our most requested plasmid, SpCas9 is still going strong as the basis for ... top requested plasmid from 2016 is an expression vector for CRISPR nuclease, SpCas9 from the Zhang Lab... of mammalian cell types. Purpose  SpCas9 This plasmid expresses Streptococcus pyogenes (Sp) Cas9 from...sequence as directed by a gRNA. 2A-Puro In this plasmid, SpCas9 is fused to the puromycin resistance gene...resistance gene enables selection of cells with the plasmid post transfection. The 2A "self cleaving" peptide... What's not in the name? Not included in this plasmid's name are some additional features. The first and...

...for your work. Species Antibody Subclass Nanobody Addgene Plasmid Rabbit N/A* TP897 pTP1183 (1x... one IgG subclass.   Express the Nanobody In Bacteria: Table 2 also has a link to the plasmid you’ll need... How to Use the Nanobody Toolbox Find Nanobody Plasmids Structural comparison As the name suggests, ...different species, or at least from different IgG subclasses, so that secondary antibodies exclusively recognize...Must use different species and/or different IgG subclasses of 1° for multi-color staining Can be expensive...limited cross-reactivity between different IgG subclasses and different species, nanobodies are useful ...require using primary antibodies from different IgG subclasses. Instead, you can pre-incubate individual primary...

...example (the top requested plasmids from 2015 and 2016 were CRISPR plasmids), but so are lentiviral vectors...reason, the top requested plasmid of 2017 is the lentivirus envelope plasmid pMD2.G from Didier Trono’...Plasmid technologies are constantly evolving, but sometimes a technology is so useful it forever enhances...’s lab! Lentivirus envelope vector pMD2.G This plasmid has been requested nearly 6,000 times (over 800...the broad usefulness of pMD2.G and VSV-G. This plasmid has been used in studies of the secretome, ubiquitin...to check out the Trono Lab website. If you have plasmids you’d like to deposit with Addgene, be sure to...Who knows, maybe yours will be the top requested plasmid of 2018! Additional Resources Quick Guide to ...

...mutation” field on the plasmid page (see plasmid 87904 for example). As many plasmids available from Addgene...blog Plasmids 101: protein expression Plasmids 101: E. coli strains for protein expression Plasmids 101...codon optimized by ordering plasmids containing them from Addgene. If a plasmid at Addgene contains a gene...

...a million plasmids would fill? Two weeks ago, Addgene edged passed our one million plasmids shared milestone...ending the day with 1,000,002 plasmids shipped. Reaching one million plasmids shared might have you wondering...how many plasmids leave Addgene headquarters every day. That would be an average of 673 plasmids each day...to packing and shipping. Every single day. 673 plasmids shipped daily may be easy to picture (see the ...celebratory lunch, it’s hard to picture one million plasmids. How much space would one million bacterial stabs...Leadership team in all their photo booth glory! #1millionplasmids On to the next million! @CarolineBME @AllostEricSite...post, we looked at how long it takes to make one plasmid and how much time Addgene has saved researchers...

...move antibiotic resistance genes between plasmids or from plasmids to the bacteria’s genome. In humans, if...elements are split into two classes based on their mechanism of transposition: Class I TEs, also called retrotransposons...rest of this article. Class I Transposable elements (TEs): Retrotransposons  Class I TEs are also known...retroviruses, like HIV, replicate. Class I TEs do not encode a transposase enzyme.  Class I TEs are considered replicative...Created in BioRender.com.   Class II TEs: DNA transposons Class II TEs are also known as DNA transposons...the key components of transposons, how they are classified, and how they are used in the lab. What is a ...retrotransposons, and Class II TEs, also called DNA transposons. DNA transposons are commonly used as tools...

...Browse our Plasmids 101 blog posts Read blog posts on plasmid cloning Read about different plasmid elements...to the backbone plasmid; a starting point could be a molar ratio of 1:2 (plasmid:insert), but this parameter...colonies screened might not contain the correct plasmid. There’s a lot that can go wrong! With all the ...cloning project, and hopefully obtain your coveted plasmid with no substantial delays. Design, design, design...spend some time outlining the construction of the plasmid and all the steps you will have to take. Decide...you chose, and verify that the sequence of the plasmid you are working with is correct. It's also best...ways you can assemble the different parts of a plasmid based on the cloning strategy you followed. While...

... sequence using BLAST2. See this blog post for tips on using BLAST to verify plasmids. Analyzer guide...SnapGene powered plasmid maps at Addgene  Three factors make our SnapGene powered plasmid maps far easier...easier for you to share plasmid information with colleagues, or explain plasmid features to a lab member... on the main plasmid page under the "Resource Information" section. Analyze your plasmid sequence For ...and “Sequence” tabs. BLAST Use the Basic Local Alignment Search Tool (BLAST) via the NCBI website ...between a given sequence and nucleotide (BLASTN) or protein (BLASTX) sequence databases. Additionally, align...over from our users: "Please, please improve your plasmid maps!" After thoughtful design, vetting, and tweaking...

... expression. Methylases involved with these processes (for example Dam and Dcm methylases) are independent...contain 3 methylases that recognize and methylate different stretches of DNA: Dam methylase adds a methyl... of DNA Dcm methylase adds a methyl group to the second cytosine of CCWGG EcoKI methylase adds a methyl...desired mutation into your plasmid of interest by PCR generates mutated plasmids with no methylation (there...Mutagenesis to Mutate Your Plasmids Resources on Addgene.org Check Out Our Plasmid Protocols Boost Your Molecular... for purposes other than cloning or confirming plasmids. It turns out that restriction enzymes are one...followed by TC. As shown in the figure below, a Dam methylase recognition site (underlined in red) overlaps ...

...Download our Plasmids 101 eBook to get all the background you need to start working with plasmids! What are...Techniques Read all of our Plasmids 101 Posts Resources on Addgene.org Browse our Plasmid Reference Pages Just...Plasmids designed to express genes in a given host cell type are generally broken down into two broad...based on the functional elements they contain. Plasmid DNA in both prokaryotic and eukaryotic systems ...Rho-dependent terminators are not usually employed in plasmid-based expression systems, so these will not be ...the end.  Nearly all common bacterial expression plasmids use Rho-independent terminators, which include...termination with polyadenlyation. Mammalian expression plasmids are primarily used to create mRNA and the commonly...

...sequence analysis, plasmid verification, and using NCBI BLAST. Tips for high-throughput plasmid sequencing If...sharing plasmids through Addgene is that we provide an added level of confidence in the plasmids we distribute...ensuring plasmid quality is to stay organized. Here at Addgene, we collect data on every plasmid we distribute...cloned into the plasmid) and any other important features that differentiate this plasmid from its predecessor...for the inserts or the full plasmids. This will enable you to quickly BLAST your sequencing result against...on the Addgene Website View our Plasmid Reference Pages Get Plasmid Backbone Information from Our Vector...distribute through our quality control processes. Every plasmid we receive is rigorously verified before becoming...

...from our repository to find our most requested plasmid in 2015. In line with all of the exciting developments... past year, we're excited to announce that the plasmid most requested from the Addgene repository in 2015...put to such great use. It’s just one of the many plasmids being distributed to and accelerating research...Knockouts Read Some Common Ways to Deliver CRISPR Plasmids to Mammalian Cells Resources on the Addgene Website...Visit our Lentiviral Guide Pages Find Lentiviral Plasmids Find CRISPR Pooled Libraries ...

...amazing plasmid technologies developed by our community of depositors. With over 40,000 plasmids avaliable...amazing new plasmid tools that have come through our doors in 2015. Do you have a favorite new plasmid tool ...system plasmids are available through Addgene and are already very popular. Use these plasmids to increase...luciferase-fluorophore pH fusion reporters, pRSETb-pHlash and pcDNA3.1+-pHlash, that use Bioluminescence Resonance Energy...Characteristics of the pHlash reporter protein include: H+ specific response cytoplasmic retention insensitivity... can visualize: Cell Structure Plasmids: Cell Structure Plasmids: ER mCherry-Sec61 beta BFP-Sec61...bound to it). Classically it has been easier to overexpress the POI-tag fusion from a plasmid or from a random...

...Addgene’s plasmids are used with a wide variety of restriction enzyme-based cloning methods. Each method...Entry DNA overhangs may be present in the original plasmid (Option 1) or added using PCR-based amplification...fragments to create TAL effector nucleases. These plasmids were designed to utilize the BsaI and BsmBI Type... a gRNA target sequence into a Cas9-containing plasmid such as pX330. This cloning strategy not only makes...makes it easy to create a single gRNA-expressing plasmid, but it can also be adapted to express multiple...create CRISPR/Cas9 constructs, assemble standard plasmids parts in different combinations, or other new ...amplification of cloned DNA as tandem multimers using class-IIS restriction enzymes. Lee JH, Skowron PM, Rutkowska...

...This post is part of our ongoing Plasmids 101 series. Plasmids 101 will provide you with an overview ...of the Plasmids 101 posts here. Now that we have covered antibiotic selection here at Plasmids 101, we...α-complementation plasmid. This way you can quickly and easily identify colonies that not only have your plasmid (antibiotic...rescued by genes supplied on the transformed plasmid. Such plasmids may contain genes that allow for the use... able to select for colonies that contain your plasmid is a great start when cloning, but how about being...being able to choose those that contain plasmid with an insert? Blue-white selection is a widely used ...far, so good, but how does this help us screen plasmids? Blue-white screening in the lab Scientists discovered...

... Hot Plasmid list. Other popular plasmids & blog posts @Addgene: The 10 Most Distributed Plasmid Technologies...inducible inactivation. Find the Robinson plasmids or the Royle plasmids and get to work! Robinson et al...designed the pOSIP (one-step integration plasmid) series of plasmids, vectors that convey both the sequence...mGRASP plasmids. Additionally, the lab recently described the use of another set of mGRASP plasmids in their... favorite plasmid technologies? Email us at [email protected]  and let us know what plasmids you would ...sgRNA libraries expand upon the CRISPR family of plasmids by offering a lentivirus-based mechanism for sgRNA...new CRISPR screening tools, see our CRISPR/Cas Plasmids: Pooled Libraries webpage or read our blog post...