...some time now. When titrating AAV by qPCR, viral DNA is amplified and monitored in real time. When the...comparing the threshold fluorescence of the viral DNA to the threshold fluorescence of a standard - a set... set of reactions with known quantities of DNA. Additionally, an AAV reference, a virus with a known titer...get right. Here at Addgene, we use a linearized plasmid standard. However, even when the standard is made... the ingredients for amplification of the target DNA. This partitioning reduces the number of PCR inhibitors... circulating Mycobacterium tuberculosis-specific DNA by droplet digital PCR for vaccine evaluation in ...

... projects, consider making you own DNA ladders with these plasmids from Penn State Need help picking the...are produced in animals instead of via recombinant DNA, they can’t be genetically tagged and instead are... a reporter and a nanobody that’s encoded on a plasmid. A nanobody is a ~150 amino acid, single domain... your own RANbodies? Great! All of the RANbody plasmids from Yamagata et al can be found here. Can’t find...Resources on Addgene.org More nanobody expression plasmids can be found here Watch this video to learn more...

...cared for your packaging cell line, prepped your DNA, transfected and harvested your lentivirus. Now it...for expression of a gene carried on the transfer plasmid or quantifying the number of viral copies that ...beneficial as they ensure that the correct transfer plasmid was used in the transfection; if preparing several... that comes from amplification of the original plasmid which can over-inflate results. A packing vector...antibiotic resistance genes carried on transfer plasmids. Target cells are transduced with serial dilutions...transduced with serial dilutions of a viral prep, genomic DNA isolated, and primers targetting viral components...In this approach, viral RNA is first converted to cDNA and then quantified using qPCR primers targeting...

...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...Lederberg's Personal Perspective: Plasmid (1952-1997) Cohen, Stanley N. “DNA Cloning: A Personal View after...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...

...scientific community obtained high-quality data for our DNA-based reagents. Not only do we extract as much information...days, we would ‘spot-check’ critical regions of plasmids deposited with us via Sanger sequencing, while...while more recently we have moved to full plasmid sequencing at our facility using Illumina MiSeq Next Generation... to a partnership with Seqera Labs.              DNA sequencing drives our bioinformatics needs   ...lab work as we grew. As the number of deposited plasmids increased, the accompanying sequencing data collection...transform the raw sequencing data into a complete plasmid sequence that would be ready for our QC team to...With our outline of individual steps to automate plasmid sequence assembly, and possible solutions for each...

... 1998). Interested in plasmids that contain the groucho gene? Check out plasmids deposited by the Shivdasani... several plasmids that contain SHH, you can find them all here! The most popular SHH plasmid however was... to see for yourself. We have one plasmid in our repository, plasmid 39624, that contains cmr.   tinman...website for plasmids that contain tinman or the mammalian homolog Nkx2.5. We also have one plasmid, Addgene...Drosophila Read our Plasmids 101 blog series Resources on Addgene.org Find plasmids for your research ...the stories behind the names. We also have many plasmids in our repository that you can use to study these..., and even humans. It binds to a specific set of DNA-binding transcription factors to repress the expression...

...bacterial in nature. All three of these methods involve DNA incorporation into the cell, but the mechanisms vary...lentivirus, adenovirus, or AAV. Then, your desired DNA to be integrated can be cloned into the vector that...typically transfected along with viral packaging plasmids into a viral production cell line, usually HEK...HEK-293 cells. When all of these viral plasmids are expressed within the production cell line, viral particles...

...transcription from the accessory plasmid (AP) encoding phage gene III. The accessory plasmid library contained all...and high DNA specificity." Nature (2018) doi: 10.1038/nature26155 PubMed PMID: 29512652 Plasmids from this...In order to bind DNA, Cas9 and other CRISPR enzymes require a short PAM sequence adjacent to the targeted...PAM flexibility. In this system, the mutagenesis plasmid (MP6) permits quick, large-scale mutation of the...Addgene.org Check out our CRISPR Guide Find CRISPR Plasmids for Mammalian Systems Learn about CRISPR History...

...can request any of over 3,000 eligible AAV cargo plasmids in the Addgene repository to be packaged in your...Request With Packaged on Request, you can find your plasmids and your viral packaging service in the same place...packaged on request process, simply navigate to your plasmid of interest. If it’s eligible for packaged on request...sequencing to confirm the identity of the packaged DNA and serotype. That way, you know you’re getting the...special processes involved; simply request your plasmid of interest in your desired serotype, place your...

...(cap) expressing plasmids is generated. Diversity is introduced into the cap plasmid library by inserting...Caltech stepped up to the challenge. Find the PHP Plasmids Here! Their Solution: Use the CREATE method to...inserting short DNA sequences that contain all possible combinations into a region of the cap gene. In this...this special CREATE cap plasmid, loxP sites flank the downstream polyA sequence. When viruses produced ...produced from this plasmid pool are injected into an animal with cell-type-specific Cre, the polyA sequence is...a cell type of interest, a scientist can isolate DNA from the infected animal and use the pre-designed... do these AAVs transduce the CNS and PNS? Both plasmids infect the PNS or the CNS much better than the...

...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...libraries containing thousands of pre-cloned gRNAs. DNA aliquots of the GeCKO v2 pooled libraries targeting...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 ...

...for sure, as is obvious from our most requested plasmid, SpCas9 is still going strong as the basis for ...experiments. So, without further ado, the most request plasmid in 2016 was... pSpCas9(BB)-2A-Puro (PX459) V2.0...      Similar to last year, the top requested plasmid from 2016 is an expression vector for CRISPR nuclease... 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...

...hepatitis B virus DNA and hepatitis C virus RNA quantification as measured with branched-DNA technology." ...when working with virus: 1. Viral vectors undergo DNA rearrangements. Depending on the composition of ...percentage of surviving cells may be the result of plasmid integration via recombination during transfection...Recombination events during integration of transfected DNA into normal human cells."Nucleic acids research 18.9...

...survivalTM, and Stbl3TM. How to transform your plasmid DNA If you decided to use commercial competent cells...if you're transforming a plasmid with ampicillin resistance.  After a plasmid is introduced to bacteria... library or an especially low concentration of plasmid), you will benefit from purchasing competent cells...negative control (plate your cells without adding plasmid) to make sure you don’t have contamination.  ... Also conduct a positive control (test a plasmid that transforms well in other competent cells) to ensure...

...contains SSB, strand displacement DNA polymerase, recombinase, and the ssDNA-FQ reporters that exhibit fluorescence...preamplification was able to detect as low as 1.2 copies DNA targets and 4.6 copies RNA targets in a 40 min incubation...sensitive to the presence of mismatches within the DNA and the positions of these mismatches. To detect ...varying between 10-50°C. Visit Addgene's COVID-19 Plasmids and Resources Many thanks to our guest bloggers...more about SHERLOCK and DETECTR Learn more about plasmids available for COVID-19 research Resources on ...in our CRISPR guide Browse our entire COVID-19 plasmid collection ...indiscriminate ssDNA cleavage activity which cleaves the supplied fluorescently labelled ssDNA substrates....

...Cas9 nuclease, so there’s no potential for aberrant DNA cleavage. Direct HIV-1 cleavage allows T-cells to...level of protein structure, CRISPR gRNAs require DNA sequence-specific binding. Patients’ HIV-1 genomes...26538064. PubMed Central PMCID: PMC4633726 Find SAM plasmids used in this publication at Addgene. Kaminski...PMC4778041. Find CRISPR and lentiviral packaging plasmids used in this publication at Addgene. Wang Z, ...Rice CM, Bhatia SN. CRISPR/Cas9 cleavage of viral DNA efficiently suppresses hepatitis B virus. Sci Rep...Addgene’s newly updated CRISPR Guide! Find CRISPR Plasmids for Your Research Browse gRNA Design Tools  ...

... 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...always to send prepped plasmid DNA for next generation sequencing. We then copy the contents of the resulting...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...

...Heidelberg University. “What is different is the plasmids you use to bring this into your target organism...stack several expression cassettes onto a single T-DNA in subsequent rounds, dramatically reducing the time...

...changes we’ve made: New CRISPR plasmid pages We’ve given our CRISPR Plasmids and Resources a fresh new design...design, sorting the plasmids we have available by category. Each plasmid page now has a helpful graphic...introduce you to the plasmid class covered on that page. To help you find popular plasmids, we’ve also added... new plasmid pages that cover some of the hottest CRISPR tools. Base Edit: Base editing plasmids allow...edits in DNA without causing double-strand breaks. Our new page shows you base editing plasmids for mammalian...revolution, Addgene has distributed over 100,000 CRISPR plasmids. But that’s not our only job - we strive to also...up with new developments (and, of course, find plasmids that will be useful to you.) With these goals...

...Addgene. I am currently working on the QC/QA team as a DNA Sequencing Intern. The application process When ... the submitted sequences were a concern to the plasmid function. The steps of the training process gave...very approachable.     Fig. 1: Examples of DNA sequencing QC work    In addition to my responsibilities...