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  1. Plasmids 101: What is a plasmid?

    Type
    Blog Post
    Published
    April 2, 2020, 6:17 p.m.
    ...cloning plasmids, expression plasmids, gene knock-down plasmids, reporter plasmids, viral plasmids, and ... of scientists and plasmid lovers - serves as an introduction to plasmids. Plasmids 101 will provide you...be asked to design, modify, or construct a plasmid. A plasmid is a small circular piece of DNA found in... components that make up a plasmid and why each is important. Our “Plasmids 101” series designed to educate...advancing the field.   What is a plasmid? At their most basic level, plasmids are small circular pieces of...cloning site. The ease of modifying plasmids and the ability of plasmids to self-replicate within a cell ...exploited for sequence verification of plasmids.   How is a plasmid constructed in the lab? Due to their...

  2. Plasmids 101: Plasmid Incompatibility

    Type
    Blog Post
    Published
    March 31, 2020, 1:15 p.m.
    ...First, why are plasmids incompatible? Plasmid incompatibility occurs when multiple plasmids within one cell...control plasmid replication of each plasmid using unique replication machinery. However, when plasmids have... one plasmid lost from the cell at a higher rate than the coresident plasmid. Asymmetric plasmid loss ...including one plasmid blocking replication of another and one plasmid outcompeting the other plasmid for replication... copy plasmid and the larger, low copy plasmid is more likely to be lost over time.  Is plasmid incompatibility...develop a plasmid-based tool that is easy to use with other plasmids.  How do you know if your plasmids are ...about plasmid cloning Resources on Addgene.org Download the Plasmids 101 eBook  Browse plasmids at Addgene...

  3. Plasmids 101: Control Plasmids

    Type
    Blog Post
    Published
    April 9, 2015, 3:29 p.m.
    ... used for plasmid-based experiments. What are control plasmids? Generally, control plasmids help to ensure...follow an example of a plasmid-based experiment, delineate its requisite control plasmids, and discuss why ...proper control plasmids would address and answer these questions. Types of control plasmids Part of planning...conditions and plasmids should produce a null effect (i.e., no phenomenon is observed). In any plasmid-based experiment... and not some other unintended factor. Control plasmids are used to minimize any effects of the non-independent...variables.  To illustrate the importance of control plasmids, we will take a step beyond discussion in the ...why these control plasmids are critical for the correct design of the experiment.  The Experiment: Knockdown...

  4. Plasmids 101: Knockout/Knock-In Plasmids

    Type
    Blog Post
    Published
    Dec. 1, 2016, 3:30 p.m.
    ...DNA designed in the lab. Specially constructed plasmids can be used to replace genes in yeast, mice, or...gene. Here, we'll describe the techniques and the plasmids used to inactivate specific genes in mammalian...mind when choosing a knockout strategy! Knockout plasmids Homologous recombination is a mechanism to accurately...randomly integrated all or large portions of the plasmid. Deliver your construct to your target cells. After...Cre-mediated recombination.   Knockout/Knock-in plasmids Gene targeting methods also make it possible to... and insert GFP into the genome, we’d create a plasmid similar to the one shown below, where the sequence...endogenous protein with GFP, as seen in blue flame plasmid OCT4-eGFP-PGK-Puro from the Jaenisch lab.   ...

  5. Plasmids 101: Optimizing Plasmid Yields

    Type
    Blog Post
    Published
    May 26, 2016, 2:30 p.m.
    ...more cells = more plasmid. Plasmids can vary in copy number, the number of plasmid copies supported within...where your plasmid yield is sub-optimal? If you are consistently getting sub-optimal plasmid yields from... optimize your plasmid prep We'll discuss the following techniques to increase plasmid yield. Keep in ...genotype, as these mutations improve plasmid stability and yield. If a plasmid is prone to recombination Use ... a strain optimized for plasmids with repeat elements, such as viral plasmids with inverted terminal repeats...co-purify with your plasmid DNA and lead to degradation of your prep. Many plasmid prep kits recommend...levels of protein, not plasmid DNA. If you need large amounts of DNA from a plasmid in a protein expression...

  6. Plasmids 101: The Wide World of Natural Plasmids

    Type
    Blog Post
    Published
    Oct. 8, 2024, 1:15 p.m.
    ...toolbox. Natural plasmids, also called environmental plasmids, are all around us! A plasmid is a small piece...colicin-producing plasmids have been referred to as “Col plasmids” or “Col factors.” Virulence plasmids: microbial...Old Solution but a New Problem Plasmids 101: What is a Plasmid? Plasmids 101: Transformation, Transduction...F. (2023). Plasmid classifications. Plasmid, 126, 102684. https://doi.org/10.1016/j.plasmid.2023.102684...., 2001) Beneficial traits of natural plasmids Natural plasmids often convey a variety of functions that...functions of natural plasmids. Created with BioRender.com. Resistance (R) plasmids: microbial defense...example of a plasmid benefiting its host. For example, cells with a penicillin resistance plasmid can grow...

  7. Plasmids 101: Broad Host Range Plasmids

    Type
    Blog Post
    Published
    April 11, 2024, 1:15 p.m.
    ...Addgene blog Plasmids 101: Origin of Replication Plasmids 101: What is a Plasmid?  Plasmids 101: Transformation...host range in plasmids? Not all plasmids were created equally. Only a handful of plasmids have been identified...broad host-range IncQ plasmids. Plasmid, 62(2), 57–70. https://doi.org/10.1016/j.plasmid.2009.05.001  Meyer...define plasmid host range, identify a few of the features that confer broad host range in plasmids in nature...recombinant vectors.  A plasmid’s host range is determined by the different hosts that a plasmid can transfer to...influence the plasmid’s ability to establish and maintain itself within host cells.  The plasmid has multiple...if a plasmid has few restriction sites, it may be able to sneak past this checkpoint.  The plasmid is ...

  8. Plasmids 101: Positive and Negative Selection for Plasmid Cloning

    Type
    Blog Post
    Published
    Aug. 22, 2019, 12:43 p.m.
    ... successfully takes up plasmid DNA. Among those that do, some now have plasmids that contain your gene...including ccdB on its destination plasmids. In the destination plasmid, the ccdB gene is flanked by recombination...selections in plasmid cloning. Additional resources on the Addgene blog Read our Plasmids 101 blog posts...purify your gene of interest, get it into your plasmid backbone, and zap the mixture of DNA into cells...gene of interest, but others will uptake plasmid backbones that re-ligated back on themselves. Therefore...strategy needs to identify cells containing the plasmid construct you’re seeking. Fortunately, there are... example, if the DNA you’re inserting into the plasmid backbone contains a kanamycin resistance cassette...

  9. Plasmids 101: Screening Strategies Used in Plasmid Cloning

    Type
    Blog Post
    Published
    Sept. 15, 2020, 1:15 p.m.
    ...once in the plasmid backbone and another enzyme that cuts within the insert. If the plasmid contains the... PCR product if the plasmid contains the insert and no PCR product if the plasmid does not contain the...resources on the Addgene blog Get all the plasmid basics in our Plasmids 101 series Find more molecular biology...If you’re cloning a plasmid, you’ll need a way to find the needle in the haystack: the one perfect clone...clone that contains the plasmid you’re looking for out of the many cells that don’t. One way to begin ... how to use positive and negative selection in plasmid cloning.  But oftentimes, a selection strategy...enough on its own to help you find your desired plasmid. In many cases, you will need a screening strategy...

  10. Plasmids 101: A Brief History of Plasmids and an Improved eBook!

    Type
    Blog Post
    Published
    Oct. 29, 2015, 2:30 p.m.
    ...tetracycline resistant plasmid, pSC101, and a newly developed kanamycin resistant plasmid, pSC102, with EcoRI...engineering, cloning, and plasmid tags, addtional content added to the "Plasmids that Glow" and "Viral Expression...Learn more about the history of plasmids and cloning: CSHL Meeting: Plasmids: History & Biology The Joshua...factors played within an organism.  So how did plasmids get their name? In 1952, Joshua Lederberg set...inheritance factors. He proposed the catch-all term “plasmid” derived as a hybrid of "cytoplasm" and "id" (Latin...” was eventually dropped and we’ve been using "plasmid" ever since! From napkins to notebooks Although...in the early 1950s, it took until the 1970s for plasmids to gain prominence in the scientific community...

  11. Plasmids 101: In-house NGS Plasmid Quality Control Now at Addgene

    Type
    Blog Post
    Published
    April 6, 2021, 12:30 p.m.
    ...every isolated plasmid sample and normalize each to fall within a 10 fold range. Plasmids that don't meet...A broader view of the whole plasmid To confirm the sequence of the plasmid, we examine three things: ...expected and do not affect plasmid function, we will update information on the plasmid’s page. Sometimes we’re...’re missing a piece of the plasmid There are regions of some plasmids that are particularly difficult ...some plasmids will not result in one complete, circular assembly. Most of the time, these plasmids are ...bottom of the plasmid page. Viral preps and pooled libraries In addition to regular plasmid sequencing,...sequence all plasmids in the pool but instead selectively sequence the variable portion of the plasmid in the...

  12. Plasmids 101: How to Name Your Plasmid in 3 Easy Steps

    Type
    Blog Post
    Published
    Feb. 13, 2014, 1 p.m.
    ...letter of a plasmid name and simply denotes that the object is a ‘plasmid’. “p” is for plasmid. pXXXXX-XXXX...information about plasmid features and tips you can use in the lab, check out more posts in our Plasmids 101 series...There are no universal rules for naming plasmids but here are some good guidelines to follow in order...people can quickly and easily identify what your plasmid contains and other important information. Tip:...Backbone name Include the empty backbone name in your plasmid name. This simple piece of information can often...important details. Once you know the backbone a plasmid is based on, you can usually derive: a) the bacterial...  Include information about the insert in your plasmid name. This is often a 3-6 letter representation...

  13. Plasmids 101: Biotinylation

    Type
    Blog Post
    Published
    Nov. 15, 2018, 1:50 p.m.
    ...biotinylating proteins with BirA. You can find plasmids that contain both AviTag and BirA at Addgene! ...

  14. Plasmid Cloning by PCR

    Type
    Blog Post
    Published
    March 29, 2016, 2:30 p.m.
    ... on All Things Plasmid with Our Plasmids 101 Series Get Tips on Verifying Your Plasmid Resources on Addgene.org...Check out Our Plasmid Reference Pages Deposit Your Completed Plasmids Find the Perfect Plasmid for Your Research...recipient plasmid (usually in the Multiple Cloning Site (MCS)), but do not cut elsewhere on the plasmid Bonus...recipient plasmid. Remember to insert your DNA in the correct orientation in the recipient plasmid by viewing... of DNA so that it can be easily cloned into a plasmid of interest. You can use similar processes to add... Interest (YGOI) for ligation into a recipient plasmid. Designing primers for PCR based cloning   ...this example). Assuming you are amplifying from plasmid DNA (rather than from genomic DNA or a cDNA library...

  15. Let There Be LITE Plasmids

    Type
    Blog Post
    Published
    Nov. 21, 2013, 3:56 p.m.
    ... (blue) light switch. Zhang’s team devised the plasmids now available in Addgene’s repository for use ...biological processes and disease states. While LITE plasmids have clear application to anyone with an interest...

  16. Plasmids 101: How to Verify Your Plasmid Using a Restriction Digest Analysis

    Type
    Blog Post
    Published
    Aug. 28, 2014, 3:34 p.m.
    ...the plasmid based on the predicted sizes and organization of different features within the plasmid. Restriction...if you don't have the full plasmid sequence. Once you have purified plasmid DNA, this method can be done...example plasmid on the right has a total size of 7.3kb, including a 1.2 kb insert. The plasmid was digested...possible plasmid conformations, with relaxed and nicked marked with asterisks (*). When the plasmid is digested... Congratulations, you have a plasmid expressing your gene of interest (YGOI) and are ready to dive into...functional experiments! Whether you’ve cloned the plasmid yourself or obtained it from a colleague down the...with the correct construct, and verify that the plasmid you received matches the expected sequence. Here...

  17. Anatomy of a Plasmid Page at Addgene

    Type
    Blog Post
    Published
    Feb. 4, 2016, 3:30 p.m.
    ... best bet is the Plasmid Map: We auto-generate a plasmid map based off of the plasmid sequence and feature...colleagues. Top of the plasmid page The ‘breadcrumb’ trail at the top of any plasmid page gives the hierarchy... to the individual plasmid page. The depositing lab page contains all the plasmids from the same lab while...contains all the plasmids deposited from the same publication. Next up, we have the plasmid name: This is...other plasmids with similar names in the collection, but you can use this name to find this plasmid from...A quicker way to find this plasmid through search is... The Addgene plasmid number: This is the unique... given to every plasmid in our catalog - if you know the catalog number of the plasmid you are searching...

  18. How to Deposit Your Plasmids with Addgene

    Type
    Blog Post
    Published
    Feb. 27, 2024, 2:15 p.m.
    ...Unpublished or pre-publication plasmids": Unpublished plasmids or plasmids that will not be published are...and entering a plasmid name,plasmid type, and plasmid description/experimental purpose (right). On the...with your plasmids, and adding all the plasmids at once increases the likelihood that the plasmids are given...easy for you to share plasmids. To achieve this goal, we will archive any plasmids you've deposited with...Spreadsheet". Start your plasmid deposit There are two options to begin your plasmid deposit. Figure 2: Choose... Choose to submit plasmid(s) online or by using a spreadsheet. The plasmid information can be entered ...than 10 plasmids but we recommend using the spreadsheet option for deposits of 10 or more plasmids.   ...

  19. Plasmids 101: Dimers and Multimers

    Type
    Blog Post
    Published
    Jan. 10, 2023, 2:15 p.m.
    ...Addgene blog:  Plasmids 101: What is a plasmid? Plasmids 101: Origin of ReplicationPlasmids 101: PCR...process known as plasmid multimerization. Plasmid Multimerization Early studies on plasmid topology recorded...ligate the plasmid, and re-transform the ligated plasmid to recover the monomeric form of the plasmid. Have... monomer of Plasmid 12259: pMD2.G (depicted on the left) is ~5.8 kb in length. The plasmid can exist on... have been utilizing full plasmid sequencing services to verify the plasmids, including long-read sequencing... the sequencing results for their plasmids, they found the plasmids were much longer than expected, and...tandem. Such plasmids are known as multimers, and they are the result of multiple plasmid copies combining...

  20. More Data for You: Find Articles Citing Addgene Plasmids

    Type
    Blog Post
    Published
    June 16, 2015, 2:15 p.m.
    ... from Addgene Plasmids 101: How to Verify Your Plasmid Plasmids 101: Control Plasmids 6 Tips for Analyzing... on our plasmid pages - links to articles citing this plasmid. Now you can learn how a plasmid has been...your deposited plasmids Increased visibility of your plasmids in the "Associated Plasmids" lists Benefits...Find other related plasmids in the "Associated Plasmids" lists Properly citing plasmids you've requested...Citing This Plasmid" list. Email us at [email protected] with the Plasmid Name, Addgene Plasmid ID #, and...'ll find the plasmids listed below the citation. An example of these "Associated Plasmids" is shown in...explore the TetO-FUW-OKSM plasmid page. Or search for your favorite plasmids and see where they've been...

  21. A Look at Addgene's QC Process

    Type
    Blog Post
    Published
    May 6, 2025, 1:15 p.m.
    ...blog Addgene's Tips for Plasmid Quality Control Plasmids 101: In-house NGS Plasmid Quality Control Now at...sequencing all incoming plasmids. If you are interested in requesting one of these plasmids, you can request ...data for each of their plasmids, from descriptions of key components of the plasmids to partial, full, or...to ensure all plasmids pass our high-quality standards and contain the expected plasmid features.  ...Once Addgene receives deposited plasmids, they undergo complete plasmid sequencing using an in-house Illumina...will compare the NGS results to the plasmid information, other plasmids in the deposit, NCBI sequence and...results and deposited sequences and plasmid information.  For each plasmid deposited at Addgene, QC scientists...

  22. Introducing Addgene's New Search - Find Plasmids More Easily For Your Research!

    Type
    Blog Post
    Published
    Dec. 3, 2019, 11:38 a.m.
    ... best plasmid for your research can be a daunting challenge. With over 80,000 deposited plasmids from ...through to the plasmid search results: More than 200 different GCaMP containing plasmids are available...search tool lets you access plasmids from journal articles. From paper to plasmid Authors regularly indicate... to include specific plasmid ID numbers. If you know the ID number for a plasmid, your straightforward...page to see the plasmids deposited by our own CSO. But what if you don’t have plasmid ID numbers, or you... Not every plasmid search will start from a publication. You may have an idea for a plasmid that would...exact plasmid you need. The ability to apply a wide range of filters isn’t new to Addgene’s plasmid search...

  23. Quickest Way to Deposit Plasmids: The Deposit Spreadsheet

    Type
    Blog Post
    Published
    March 8, 2017, 3:15 p.m.
    ...deposits with more than 10 plasmids is to use our deposit spreadsheet?  Our plasmid deposit process can be... Send plasmid information to Addgene Material Transfer Agreement (MTA) approval  Send plasmids to Addgene...spreadsheet and copy and paste your plasmid data directly into our file, one plasmid per row, as outlined by the... Data Field Description Plasmid Name Use a descriptive plasmid name and match the name as it ...application of the plasmid and will help a scientist outside your field understand if this plasmid will work for...possible, we propagate plasmids in the standard cloning strain DH5α. For plasmids with highly repetitive...lentiviral, retroviral, and AAV plasmids, choose the NEB Stable strain. For plasmids containing the ccdB gene...

  24. Increasing Your Work's Visibility with Addgene: Citation, Search, and Collections

    Type
    Blog Post
    Published
    March 10, 2016, 3:30 p.m.
    ...curating plasmid collections. Addgene's "How to cite this plasmid" section Anytime you use a plasmid from ...process, we explain how to cite plasmids from Addgene right on our plasmid pages and provide a citation ...comparing two plasmids, we recommend reviewing the cited papers to determine whether the plasmid has been ...collection or if you think a particular plasmid or type of plasmid is missing from our collections pages...spotlight some of the plasmids that have come through our doors in our “Hot Plasmids” section. We’ve also... added the Plasmid of the Day feature to our home page - each day we showcase a new plasmid that you might...variety of plasmids in the repository. And last, but certainly not least, we often feature new plasmid technologies...

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

    Type
    Blog Post
    Published
    May 19, 2020, 1:15 p.m.
    ...What SARS-CoV-2 plasmids from Ginkgo Bioworks are available for request? Currently, plasmids containing all...available. Plasmids are continually added as they complete QC and you can find all available plasmids listed...to fresh data! These same plasmids are then added into our normal full plasmid NGS queue in order to bring...pipeline. For the first ~50 plasmids we perform our standard full plasmid sequencing by NGS. Addgene’s...on Addgene.org Find all plasmids from Ginkgo Bioworks  Find COVID-19 plasmids for your research Browse... to synthesize and share over 1,850 SARS-CoV-2 plasmids to help academic and industrial labs accelerate...confirmed the expected features and sequences in each plasmid, none of these constructs have been functionally...

  26. Unique and persistent IDs for improved reproducibility: Addgene now supports RRIDs and compact identifiers for all plasmids

    Type
    Blog Post
    Published
    Nov. 20, 2018, 8:17 p.m.
    ...materials that we use in our labs, including plasmids. All plasmids at Addgene have unique ID numbers associated...identify a plasmid available through Addgene and find verified information about each plasmid item. Given...associated plasmid page on the Addgene website and find specific information about the plasmid: name, description...description, how the plasmid was generated, validated sequence information, and more. The plasmid page also helps...publications that have used the same plasmid through the “Articles Citing this Plasmid” section. Having IDs linked...specific plasmids used. RRIDs and resolvable compact identifiers now exist for all Addgene plasmids! In an...our example plasmid, lenti-Cas9-VQR-Blast. If you scroll down to the bottom of this plasmid’s page you will...

  27. Three Tips for Preventing Viral Plasmid Recombination in Your Samples

    Type
    Blog Post
    Published
    Sept. 14, 2021, 1:15 p.m.
    ...viral vectors and plasmids with repeating sequences. Repeating sequences within a plasmid can undergo intramolecular...contain the full length plasmid.   Figure 1: The full lentiviral plasmid is expected to be >12 kb...in my samples? Plasmid recombination can affect viral vectors (LTRs and ITRs), plasmids with multiple ...other plasmids that contain repeating elements. Knowing how to amplify recombination-prone plasmids and ... length plasmid. If you find that your bacterial cultures contain a mix of full length plasmid and the...smaller, recombined plasmid around 1.5 kb. When cut with BamHI, the expected linear plasmid is present, but...get to see everything that can go wrong with a plasmid. One of the most frequent problems that I help ...

  28. New Podcast Segment: Hot Plasmids

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    Blog Post
    Published
    May 17, 2017, 3:37 p.m.
    ...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...

  29. 15 Years of Addgene: The Top 15 Plasmids

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    Blog Post
    Published
    Jan. 8, 2019, 1:54 p.m.
    ...and an envelope expressing plasmid, such as our second most requested plasmid of all time, pMD2.G. Find...wide range of applications. This plasmid was the top requested plasmid of 2017. Find pMD2.G. pSpCas9(BB... top requested plasmid from 2016. You may notice the “V2.0” at the end of this plasmid which indicates...prior version of this plasmid. Indeed the first version is the 15th most popular plasmid of all time, but ...envelope expressing plasmid pMD2.G. Find pMDLg/pRPE. scramble shRNA - This plasmid from David Sabatini....1 protocol. Pro-tip: this plasmid grows more slowly than standard plasmids. Find pLKO.1 - TRC cloning...15 years of plasmid sharing has certainly taken us on many adventures...From moving office locations ...

  30. 500,000 Plasmids Shipped and Counting!

    Type
    Blog Post
    Published
    March 26, 2015, 3:24 p.m.
    ... here at Addgene – 500,000 plasmids shipped! That's a 1/2 million plasmids sent to happy scientists around... you all the plasmids you need. We're focused on our mission – documenting your plasmids, storing them...500,000 plasmids go? Far and wide – that's where! Check out the map below which depicts where plasmids have...Sidney Street. Then the plasmids travel to labs around the world. We've shipped plasmids to more than 90,000...providing quality control and customer support, making plasmids easy for you to find and request in our online...shipping them to you quickly.   So, what does plasmid shipping look like at Addgene? Here's an inside... workings in Addgene's lab. We ship individual plasmids as transformed bacteria in stab culture format...

  31. Plasmids 101: Walkthrough of Addgene’s Snapgene-Powered Quality Control Process

    Type
    Blog Post
    Published
    Feb. 1, 2018, 3:07 p.m.
    ... we use SnapGene for plasmid quality control are: To identify important plasmid features To align NGS...design experiments for further plasmid verification Identifying plasmid features While our quality control...control process for incoming plasmids can vary based on the contents of a plasmid, the first step is always...send the plasmid off for Sanger sequencing.     If a repetitive region is present in the plasmid, we might...sequencing results for plasmids in the repository at a glance. Our Snapgene-powered plasmid maps and sequences...the Addgene Blog How to verify your plasmid Check out our Plasmids 101 featured topic page Additioinal...results to perform quality control on deposited plasmids. We’ve also talked about how our new Snapgene ...

  32. The Blue Flame Award: Celebrating Addgene's Most Requested Depositors

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    Blog Post
    Published
    Nov. 14, 2016, 3:30 p.m.
    ... Requested Plasmid in 2015 Find Hot Plasmid Technologies Write about Your Favorite Plasmid Technology ... indicates that a plasmid has been requested 20+ times   A red flame means a plasmid has been requested...answering plasmid requests can be a time-consuming chore for labs that have popular plasmids but lack ...All plasmids deposited at Addgene enhance scientific sharing worldwide but a select few are particularly...particularly special. Over 1000 of our 60,000+ deposited plasmids have been requested more than 100 times each, ...to sharing epitomized by the creators of these plasmids with the first annual Blue Flame Awards. What'...scientists around the world choose to share their plasmids through our repository since we know biological...

  33. Site Directed Mutagenesis by PCR

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    Blog Post
    Published
    Aug. 2, 2016, 2:30 p.m.
    ...mutagenesis of Plasmids. A) Generation of mutagenized plasmids. PCR primers (green) amplify the plasmid template...with the nuclease-resistant nicked plasmid (the PCR product). Plasmids are isolated from the resulting colonies...the final circular plasmid product pictured here. B) Screening of recovered plasmids for the mutation of...Smaller plasmids (~3 kb) are generally more efficiently amplified than larger constructs, but plasmids as ...DpnI) to eliminate the parent plasmid from the PCR products, the plasmid template should be isolated from...have taken up the mutagenized plasmid. Note that any residual parental plasmid (usually from incomplete DpnI...additional restriction site into your plasmid, then, when the plasmid is digested with the appropriate restriction...

  34. Bacterial Toxin-antitoxin Systems as Molecular Biology Tools

    Type
    Blog Post
    Published
    Nov. 1, 2018, 12:35 p.m.
    ...cells to retain plasmids - if a cell does not inherit plasmid due to improper plasmid partitioning, it...unstabilized plasmids were retained in less than 10% of cells while plasmids containing the plasmid addiction...ensure that plasmids are passed down through generations (Figure 1). These so called “plasmid addiction ...only survive if they retain the plasmid. Since their discovery, plasmid addiction systems have been used...found that if a mini-F plasmid carried a specific 700 bp segment of DNA, the plasmid was propagated better...selection in one plasmid, they were able to create an efficient cloning tool. The plasmid pKSCTMF contains...restriction-modification systems onto plasmids allowed cell populations to retain more of these plasmids as compared to those...

  35. Twenty Years of Addgene Sharing: CRISPR

    Type
    Blog Post
    Published
    April 18, 2024, 1:15 p.m.
    ... (A) Cumulative CRISPR plasmids shipped by year. (B) Cumulative CRISPR plasmids deposited with Addgene...,000 CRISPR plasmids in our repository, and we have shared almost 300,000 CRISPR plasmids and 12,000 CRISPR...these CRISPR plasmids (and libraries)? We wanted to know, so we classified CRISPR plasmids containing a...CRISPR plasmids requested each year by year of deposit (Figure 3), we saw that once CRISPR plasmids become... CRISPR plasmids seem to never truly go out of style. Instead, an almost even mix of plasmids (by age)...Total CRISPR plasmids deposited by country; square size is proportional to number of plasmids deposited ...Total CRISPR plasmids requested by country; square size is proportional to the number of plasmids requested...

  36. The time and cost required to make a plasmid

    Type
    Blog Post
    Published
    March 23, 2023, 1:15 p.m.
    ...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 ...

  37. The 10 Most Distributed Plasmid Technologies in Addgene's First 10 Years

    Type
    Blog Post
    Published
    Jan. 8, 2014, 3:10 p.m.
    ...cloning vector. Lentiviral Plasmids -  If you're familiar with the pLKO plasmids you'll note that those are...Actually, these two plasmids from Didier Trono's lab are our two most popular plasmids of the past 10 years...influx of plasmids for reprogramming differentiated cells into pluripotent stem cells. Plasmids from Shinya... have 19 different ubiquitin plasmids in the repository. 100,000 Plasmids Shipped! - By 2010, Addgene ...our stride. We shipped our 100,000th plasmid. A variety of plasmids continued to be popular years after...with all the new plasmids. So far, we have distributed over 10,000 CRISPR/Cas9 plasmids to more than 3,...10 years of plasmid sharing  Interested in more facts about Addgene's 10 years of plasmid sharing? Check...

  38. CRISPR 101: Multiplex Expression of gRNAs

    Type
    Blog Post
    Published
    Sept. 10, 2020, 11:45 a.m.
    ...multiple gRNAs on the same plasmid, you’ll make sure that each cell that gets the plasmid contains all of the...rectangles) are cloned into various plasmids using oligonucleotides. These plasmids contain Type IIS restriction...different purposes.   Gersbach Lab multiplexing plasmids: This plasmid set allows you to express 2-4 gRNAs, with...assembled to produce a plasmid encoding the gRNAs and Cas9. As with the Gersbach lab plasmids, multiple Cas9 ...Golden Gate/Gibson Assembly Multiplexing Plasmids: These plasmids allow you to assemble 2-4 gRNAs through...Golden Gate/Gibson Assembly Multiplexing Plasmids: These plasmids can be used to express up to 8 gRNAs after...Yang Lab Single Transcript Multiplexing Plasmids: These plasmids are similar to the Csy4 polycistronic ...

  39. Plasmids 101: Stringent Regulation of Replication

    Type
    Blog Post
    Published
    June 20, 2024, 1:15 p.m.
    ...replication. In general, these plasmids tend to be low copy number. Conversely, plasmids that can initiate DNA ...growth against sustainability. Plasmid DNA replication and regulation Plasmids must also strike a balance:...a given plasmid has been reached, negative feedback circuits reduce replication of the plasmid to balance...balance the total plasmid number.  Let’s consider pSC101. This plasmid is stringently controlled and has...promotes initiation at the plasmid ori, after which replication of the plasmid proceeds (and terminates)...tool to stringently regulate plasmid replication. For example, the plasmid ColIb-P9 encodes the protein...All plasmids rely on their host cell's replication machinery in order to replicate—but not always to ...

  40. Plasmids 101: Origin of Replication

    Type
    Blog Post
    Published
    Nov. 11, 2020, 2:23 p.m.
    ...consider your plasmid's compatibility with one or more other plasmids. Generally speaking, plasmids with the...enabling a plasmid to reproduce itself as it must to survive within cells. The replicons of plasmids are generally...cell. Figure 1: A plasmid map showing the standard features of a plasmid. So, how do you choose...have only one plasmid type in your cells at a time? Do you want to make a lot of your plasmid? Is the gene...avoid plasmid incompatibility In other words, the best choice of ori depends on how many plasmid copies...a rule, plasmids from the same group should not be co-transformed, so if you need two plasmids for an ...purify your plasmid DNA: The insert Bacteria tend to maintain fewer copies of plasmids if they contain...
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