... 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...

...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...

...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...

... biology tools for the lab. Natural plasmids Plasmids found in nature often give their hosts beneficial...environments. Plasmids derived directly from the environment are sometimes called ‘natural’ plasmids, to distinguish...natural plasmids enable horizontal gene transfer, allowing beneficial traits encoded by these plasmids to ...been derived from the natural yeast 2-micron plasmid. Plasmid transfer outside the lab In the lab, we use...transfection to put plasmids into cells. In nature, the two main routes of plasmid transfer are conjugation...Benefits of natural plasmids What actually constitutes a “beneficial trait” encoded by a plasmid? In addition... be passed around on plasmids like a “get out of jail free card.” These plasmids were historically called...

...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.   ...

...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...

...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 ...

...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...

... 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...

...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...

...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...

...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...

... 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...

...biotinylating proteins with BirA. You can find plasmids that contain both AviTag and BirA at Addgene! ...

... (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...

...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...

... 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...

...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...

...Pre-Publication or Unpublished Plasmids”: Unpublished plasmids or plasmids that will not be published are... to add plasmids to your article: Figure 3: Add plasmids to the article by entering a plasmid name, plasmid...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... submit published or unpublished plasmid(s) online or submit plasmid(s) using a spreadsheet. Fields marked...Submit Plasmids from a Published Article”: This option allows you to quickly associate your plasmids with... final plasmid pages and scientists will be asked to cite this article when using your plasmids. You ...

... 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...