...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...quantity inside a cell Plasmids 101: Green Fluorescent Protein (GFP) Plasmids 101: Luciferase Monitor...components that make up a plasmid and why each is important. Our “Plasmids 101” series designed to educate...specific plasmid element that you would like answered or any topic suggestions for our Plasmids 101 series...be asked to design, modify, or construct a plasmid. A plasmid is a small circular piece of DNA found in...advancing the field.   What is a plasmid? At their most basic level, plasmids are small circular pieces of...

...about plasmid cloning Resources on Addgene.org Download the Plasmids 101 eBook  Browse plasmids at Addgene...First, why are plasmids incompatible? Plasmid incompatibility occurs when multiple plasmids within one cell...Replication? See our Plasmids 101: Origin of Replication blog).  The replication problem Plasmids with the same...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...

... 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...Promoter Region – Let's Go! Browse All of Our Plasmids 101 Posts       ... 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 ...

... An Old Solution but a New Problem Plasmids 101: What is a Plasmid? Resources on Addgene.org Addgene's... 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...

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

...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...Addgene Blog Visit our Plasmids 101 Topic Page Learn How to Verify Your Plasmid Learn about Common Lab...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...

...applications in our Antibodies 101 blog series New to antibodies? Read our Antibodies 101: Introduction to Antibodies...interest endogenously and transfect them with a plasmid encoding your protein of interest. Then you should...target protein in a Western blot. You acquire a plasmid encoding your protein of interest, known as Gene... In parallel, you transfect cells with another plasmid not encoding this protein, which we will call Negative...band from the cells transfected with the other plasmid. Your antibody works! Multiple (Independent) Antibodies...Antibodies blog post!  Resources on Addgene.org Find plasmids encoding antibodies at Addgene Stay tuned for ...Designing a recombinant validation experiment? Find plasmids here!  References Baker M (2015) Reproducibility...

...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...

...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...

...More: Plasmids 101: Antibiotic Resistance Genes Plasmids 101: Origin of Replication All Plasmids 101 Articles...Although plasmids do not naturally exist in mammals, scientists can still reap the benefits of plasmid-based...their genomes. Unlike plasmids used for expression in yeast and bacteria, plasmids used for stable transfections...Expression Plasmids? Check Out Addgene's Empty Backbone Resource: Choosing Your Perfect Plasmid Backbone...origin of replication (ORI) will not allow for plasmid replication in mammalian cells, for example, and...In this blog post we will discuss how mammalian plasmids differ from their bacterial counterparts, including...transfected cells. Before getting into the mammalian plasmid components, it may be useful to describe the means...

...How to Verify Your Plasmid Try Out Golden Gate or SLIC Cloning Read Other Plasmids 101 Posts Resources ...covalently binds the plasmid to the new fragment thereby generating a complete, circular plasmid that can be easily...restriction digests for your insert (or donor plasmid) and plasmid backbone. Because you lose some DNA during...insert and 1μg of plasmid backbone. It is also critical that as much of the backbone plasmid as possible be... of the backbone plasmid (see below). You should treat your digested backbone plasmid with a phosphatase...ligations for each plasmid you are trying to create, with varying ratios of recipient plasmid to insert. It...“recipient plasmid : insert ratio” in the ligation reaction. 5. Purify the finished plasmid   Once it...

...our first few Plasmids 101 posts, we focused mainly on the elements required for plasmid maintenence within...types of yeast plasmids are defined below: Yeast Integrating plasmids (YIp): These plasmids lack an ORI...not only the number of plasmids found in each cell, but also whether the plasmid gets integrated into the...integration. Yeast Episomal plasmids (YEp): These are most similar to bacterial plasmids and are considered “...simple gene replacement/mutation. Yeast expression plasmids used in the lab typically contain all the necessary...how the regulation of bacterial ORIs determines plasmid copy number within the bacterial cell. Similary...included within a yeast vector determine how the plasmid is replicated and maintained within the yeast cell...

... one-stop reference guide for plasmids. We've combined our Plasmids 101 blog posts and additional resources...desktop. The Plasmids 101 eBook is designed to educate all levels of scientists and plasmid lovers and ...May 11.) PubMed.   Read more Plasmids 101 posts or browse Addgene's Plasmid Guide for more molecular biology...with a plasmid that already has the multicistronic element and reporter cloned in. In these plasmids you ...interest. Some of the plasmids listed in the table above (and their related plasmids) are designed to have...for easy detection.5  Just released: Addgene's Plasmids 101 eBook And now for some exciting news... You ... an introduction to plasmids, allowing you to spend less time researching plasmid basic features and spend...

...Deletions in a Plasmids Learn How to Verify Your Plasmid Resources on Addgene.org Find More Plasmid Cloning...  Proc Natl Acad Sci U S A.  1991 Nov 15;88(22):10104-8. PubMed PMID: 1658796. PubMed Central PMCID: PMC52876...

...the correct orientation or even if it’s in your plasmid backbone. Backbone-specific primers: A second...orientation of your insert or if the insert is in your plasmid Have to make a new primer pair for each insert ...about the size of the insert and if it's in the plasmid Can be used to screen clones that were created ...created with the same backbone because primers are plasmid dependent Doesn't provide information about the...insert Tells you if the insert is present in the plasmid Great for blunt end cloning You have to make...thermocycling program. One key difference is the plasmid DNA must be released from the bacteria in order... just need to lyse the bacteria to release the plasmid DNA by either briefly boiling the sample before...

...In a previous post from our Plasmids 101 series, we learned how the Cre-loxP recombination system can... plasmids 101... even in the presence of Cre recombinase. This plasmid now specifically drives expression of mCherry instead...2286602. PubMed Central PMCID: PMC3406316. Find FLEx plasmids from these Addgene depositors: Ed Boyden Bernardo... plasmids plasmid... plasmid plasmids... flex plasmid...

...Tet-Off system.  For more information on Addgene Tet plasmids, see our Tetracycline resource page. Cumate Switch...20697123. PubMed Central PMCID: PMC2942869. Find plasmids from this publication at Addgene. del Valle Rodriguez...20434990. PubMed Central PMCID: PMC2883883. Find plasmids from this publication at Addgene. Wei, Xing, ...22406855. PubMed Central PMCID: PMC3846601. Find plasmids from this publication at Addgene. Riabinina, ...25581800. PubMed Central PMCID: PMC4344399. Find plasmids from this publication at Addgene. Riabinina O...Hill, C. et al. A LexAop > UAS > QUAS trimeric plasmid to generate inducible and interconvertible Drosophila...expression. Genes Dev. 1988 Jun;2(6):718-29. doi: 10.1101/gad.2.6.718. PMID: 2843425.Takara Bio. Tet-On and...

...blog Plasmids 101: Protein Tags Plasmids for Endogenous Gene Tagging in Human Cells Antibodies 101: Epitope...Addgene.org CRISPR Plasmids – Protein Tagging CRISPaint Gene Tagging Plasmids 101 eBook Resources on...cloned in-frame to the protein of interest for plasmid-based expression systems. Degron tags are one of...

...’s DAD Interview with Nicola Patron on MoClo Plasmids 101: Golden Gate Cloning ...As a plasmid repository, we are no strangers to cloning. If you’re reading this, then chances are you...