... as a donor plasmid containing the transposon, along with a helper plasmid that expresses the transposase...lingering expression of reprogramming factors. Non-replicating Plasmids: Generating iPSCs with plasmid-based...plasmid-based expression requires serial transfection of 1 or 2 plasmids that express the reprogramming factors...mutagenesis, transgene reactivation, residual expression Lentivirus Integrating High Low Efficient and Stable...reprogramming. Additionally, lingering expression or re-expression of viral transgenes in iPSC-derived cells...cassette that expresses multiple reprogramming factors driven by one promoter. While lentivirus has similar... generating iPSCs by plasmid transfection where 2 of 11 clones tested had plasmid integration. Another...

... Plasmid Collections Plasmids for Stem Cell Research Plasmids for Stem Cell Research...Reprogramming Plasmids Differentiation and Transdifferentiation Plasmids Stem Cell Research Plasmids Stem cells...utilize plasmids to increase expression of key factors. The tables below will help you find plasmids available...information page. Reprogramming Plasmids Browse the table below for plasmids used to create induced pluripotent...Method Species Description Article PI Lentivirus Human Expression of human Oct4, Sox2, Klf4, Myc, and Brd3R...Commun. 2018 Jul 6;9(1):2643. Otonkoski Lentivirus Human Expression of human Sox2, Nanog, Oct4, and Lin28...318(5858):1917-20. Thomson Lentivirus Human Doxycycline-inducible expression of human Oct4, Sox2, Klf4,...

...CRISPR plasmids available at Addgene. We will still periodically focus on specific CRISPR plasmid tools...CRISPR guide Find CRISPR plasmids by function Find recently deposited plasmids ...of bacteria that can cause disease in humans. Expression of this system is tunable via a rhamnose-inducible...lentiviral capsid-based bionanoparticle Transient expression of CRISPR/Cas9 in cells has advantages like reduced...packaging Cas9/gRNA ribonucleoproteins for transient expression. They replaced the tetraloop of the sgRNA scaffold... aptamer binding protein for packaging into a lentivirus-like particle. Read the paper in Nucleic Acids...Acids Research Find the gRNA and packaging plasmids   Cytosine deaminase for RNA editing Feng Zhang’s ...

...packaging plasmids. The cells need three different plasmids to produce AAV: 1) the RepCap plasmid which provides...AAV plasmid standard and AAV transfer plasmids. It’s easy to contaminate a workspace with plasmid, not...protein shell, or capsid; 2) the pHelper plasmid which expresses adenovirus genes which help mediate AAV...going to lie, AAV transfer plasmids are kind of a pain to work with. These plasmids contain ITRs which are...AAV viral replication; and 3) the transfer plasmid which encodes a transgene of interest that’s packaged...structures that are prone to deletion from the transfer plasmid. The good news is there’s a few simple ways to ...multiple bacterial cultures of your AAV transfer plasmid at 30 °C instead of 37 °C and then screen for ITR...

...the transfer plasmid, a plasmid encoding Rep and serotype-specific Cap genes, and a plasmid encoding adenoviral... coli . In addition, plasmids are typically prepared using endotoxin-free plasmid purification kits. Resources...viral production methods and quality control for lentivirus and AAV. Viral...AAV Production Titer Quality Control Resources Lentivirus Production Titer Quality Control Resources Overview...contamination is minimized by using an endotoxin-free plasmid purification protocol. To minimize the immunogenic...vectors are tested in vitro and in vivo for gene expression and/or function. These data are sometimes reported...mL, or both. Two days later, Cre-dependent GFP expression was detected with direct fluorescence. GFP was...

...from Rudolf Jaenisch’s lab in plasmid form or as ready-to-use lentivirus. Figure 2: dCas9-Tet1 ... are an additional layer of control over gene expression that go beyond genomic sequence. Dysregulation...modifications are correlated with alterations in gene expression, but causality and mechanisms remain unclear....that help package DNA, were associated with gene expression or silencing. To alter DNA modifications, researchers...Charles Gersbach’s lab has deposited mammalian expression constructs including pcDNA-dCas9-p300 Core and...well as pLV-dCas9-p300-P2A-PuroR for lentiviral expression. Figure 1: dCas9-p300 adds H3K27ac marks...cytosine demethylation in mammalian cells. This plasmid is used with pcDNA3.1-MS2-Tet1-CD to decrease methylation...

... the lentiviruses (like HIV-1) — that have been developed into lentiviral vectors. Lentiviruses are sometimes...instead get expression of the sensor, marker, or whatever other tool you chose to express. In the case...that the virus must have its structural proteins expressed from a reverse-transcribed piece of DNA called...is done, and the transgenic genome will simply express from whatever promoters were included in the vector...And there you have it, the full “life cycle” of expression of selfish, lazy genes of a non-living viral ...Addgene's Lentiviral Guide Most popular lentiviral plasmids γ-Retrovirus Guide References Cullen, B. R. (...

...vectors and an envelope expressing plasmid, such as our second most requested plasmid of all time, pMD2.G.... the envelope expressing plasmid pMD2.G. Find pMDLg/pRPE. scramble shRNA - This plasmid from David Sabatini...(PX461) expresses both a Cas9 protein and a gRNA. But instead of Cas9, this plasmid expresses Cas9n (a...generation lentiviral packaging plasmid from the Didier Trono lab that expresses the HIV gag, pol, rev, and...Didier Trono lab is pMD2.G, a VSV-G envelope expressing plasmid. Envelope proteins determine which types ...wide range of applications. This plasmid was the top requested plasmid of 2017. Find pMD2.G. pSpCas9(BB...BB)-2A-GFP (PX458) - This plasmid from the Zhang lab expresses Cas9 from Streptococcus pyogenes with an...

...permanently induce expression? Check out our Retro expression vector and Lenti expression vector. Turn it...ablate protein expression. If you need the ability to control when you turn off expression for good, the...inducible expression (more on this below) will both achieve temporary repression of protein expression. Addgene... a gene (cDNA) can be done by either plasmid or virus. A plasmid carrying your cDNA can be targeted for...appropriate promoter to express your cDNA, depending on the level of expression you would like. Check out...choosing. Viral expression is another mechanism to integrate your choice cDNA. Both lentivirus and retrovirus.... Whether you want to express or repress, good luck on your protein expression journey! Resources and...

... to find plasmids that can be used to engineer various types of viruses. Lentivirus Plasmids | Guide Lentiviruses...enabling long term transgene expression. Overview of Viral Plasmids While plasmids are typically used to encode...for select plasmids are now available through Addgene, learn more here . Browse Viral Plasmids by Type Visit...direct high but transient transgene expression. γ-Retrovirus Plasmids | Guide γ-retroviruses, commonly referred...organism. Figure 1. Using viral plasmids to generate virus. A) Viral plasmids have been engineered such that...genomes have been adapted into a plasmid-based technology, such that plasmids can be used to create viruses...researchers can introduce plasmids to a host to generate virus. Furthermore, these plasmids can be modified to...

...μg pLKO.1 shRNA plasmid 750 ng psPAX2 packaging plasmid 250 ng pMD2.G envelope plasmid to 20 μl serum-...to vary the ratio of shRNA plasmid, packaging plasmid, and envelope plasmid to obtain the ratio that gives...drives expression of puromycin. Puro R Puromycin resistance gene for selection of pLKO.1 plasmid in mammalian...shRNA. psPAX2 Packaging plasmid for producing viral particles. pMD2.G Envelope plasmid for producing viral... also be used with packaging plasmid pCMV-dR8.2 dvpr and envelope plasmid pCMV-VSVG from Robert Weinberg...into the pLKO.1 vector, producing a final plasmid that expresses the shRNA of interest. C.1 Recommended ... transient knockdown of protein expression, you may transfect plasmid DNA directly into the target cells...

...cell page Find plasmids for reprogramming and differentiation Check out our lentivirus guide Sign up to...), now allows scientists to identify the gene expression patterns unique to the few cells that successfully...Unfortunately scRNA-seq only captures the gene expression of cells at a single time point, making it difficult...difficult to investigate how early expression patterns influence reprogramming outcomes. CellTag from ...single-cell level, thus linking ancestral gene expression patterns with reprogramming outcomes (Biddy et...CellTags were detected over a 10-week period. Their expression decreased over time but they were not silenced...MEFs were reprogrammed into iEPs by retroviral expression of transcription factors Foxa1 and Hnf4α, a process...

...and lentiviral expression vectors (Table 1). The four systems Name Function Plasmid Transduction...deposit also includes pXPR_071, a plasmid that leads to higher titer lentivirus preparations than the first ...2019b). CHIME works through transduction of Cas9-expressing (Rosa26-Cas9) hematopoietic stem cells with a...first generation CHIME (pXPR_053). Find X-CHIME plasmids here!   Marty LaFleur is a Postdoctoral Fellow in...

...Addgene.org Plasmid Pooled Libraries Pooled library amplification Zhang Lab CRISPR Plasmids The Human TFome...factors (TFs) act as the gate keepers for gene expression, turning transcription on and off by binding ...binding proximal to their target genes. Since gene expression patterns determine everything from stem cell differentiation...(aka the MORF Collection) and an accompanying expression atlas, including over 3,500 human TF isoforms...TF library, with each cell receiving only 1 TF expression construct. Cells were cultured following a differentiation...from Joung et al.)    This atlas linked TF expression to differentiation outcomes as well as global...reference cell types generated by the human fetal expression atlas. Computational analysis was used to infer...

...transfer vector for in-vivo expression and plasmids optimized for dCas9-VPR expression in drosophila and yeast...Cas9 nickase, and Cas9 activator expressing plasmids that can co-express 1-8 gRNAs. Ordon et al., 2016 tested...Neurosci 2016. PubMed PMID: 26950006 Plasmid ID Plasmid Name Plasmid Type 74309 pcDNA3.0-TRPV4-p2A-ferritin-p2A-mCherry...highlight a subset of the new plasmids in the repository through our hot plasmids articles. These brief articles...features and applications of a partiular plasmid or set of plasmids. We hope that these articles make it ... you to find and use the plasmids you need. You can find all the hot plasmids from 2016 below. With over...over 45,000 plasmids, we can't write posts for every great plasmid that comes into the repository, but...

... have found that expression occurs quickly and results in long and stable expression (up to 9 months in...the onset of gene expression. However, it has yet to be determined what gene expression looks like at 3 ...facilities (Biosafety Level-1). Other possible tools, lentiviruses for example, are not approved for use in vivo...Response: Compared to other viral vectors (e.g. Lentivirus, Adenovirus) AAVs cause the least amount of immune...AAV2 can transduce many tissue types, it has low expression in several brain regions where other serotypes... under study. Many AAVs will show significant expression in a variety of cell types, but you should try...transport desirable or not desirable? If diffuse gene expression is tolerated, then axonal transport is fine and...

...Plasmids designed to express genes in a given host cell type are generally broken down into two broad...Download our Plasmids 101 eBook to get all the background you need to start working with plasmids! What are...terminators are not usually employed in plasmid-based expression systems, so these will not be detailed...at 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...Polyadenylation Signal Substantially Increases Expression Levels of Lentivirus-Delivered Transgenes but Has the Potential...Promoter Controls Gene Expression Learn about New Cloning Techniques Read all of our Plasmids 101 Posts Resources...

...donor plasmid or single-stranded DNA oligonucleotide, so you will need to make sure your expression system...included with each expression system as well as suitable applications. Expression System Components ...System Application Mammalian expression vector Promoter driving Cas9 expression can be constitutive or inducible...or electroporation. Transient expression of CRISPR components, expression decreases as RNA is degraded ...using cationic lipids. Transient expression of CRISPR components, expression decreases as gRNA and Cas9 protein... provide an overview of some CRISPR mammalian expression systems, the typical applications for each, and...gene? Do you want to activate or repress gene expression? Are you trying to create a single point mutation...

...promoter, which is used to drive the expression of the array. After expression, the individual gRNAs are released...demonstrated that the tRNA-gRNA array can drive the expression of itself, without using any additional promoter...deliveries via adeno-associated virus (AAV) and lentivirus, demonstrating simultaneous editing of genes ..., that rely on an upstream lengthy promoter to express the gRNA array, DAP arrays use tRNA itself, as ...tRNA-gRNA arrays, which is scalable and efficient in plasmid and viral vectors. In applications when repetitive...manual and online design examples of using Addgene plasmids hCtRNA_FT (#186715) and hCtRNA_VT (#186716).   ...resources on the Addgene blog CRISPR 101: Multiplex expression of gRNAs Four Base Editing Reporters to Monitor...

...Adenoviral Guide Viral Vector Plasmids and Resources Addgene Blog: Your Lentiviral Plasmid FAQs Biosafety Resource...reporter plasmids to test whether or not you've efficiently generated infectious virus, plasmids expressing...-EF1-DIO-copGFP 72253 Expresses copGFP under EF-1 promoter when Cre is expressed by another vector pCDH-EF1...pCDH-EF1-DIO-tdTomato 72254 Expresses tdTomato under EF-1 promoter when Cre is expressed by another vector pCDH-CB-iCre-P2A-tdTomato-T2A-Puro...pCDH-CB-copGFP-T2A-iCre 72256 Expresses copGFP and iCre under from the CB promoter pCDH-CB-iCre 72257 Express iCre under ...pCDH-CB-FLPe-P2A-tdTomato 72259 Expresses FLPe and tdTomato from the CB promoter pCDH-EF1-Fon-copGFP 72260 Expresses copGFP from...promoter when FLP is expressed by another vector pCDH-EF1-Fon-tdTomato 72261 Expresses tdTomato from the ...

...applications Gene expression control Researchers are always wanting to manipulate gene expression – turn it up...interest, turning expression off permanently. Viral vectors can also be used to express proteins. Viral... the genome of the target cell or transiently expressed. It’s important to note that for all the applications...is more than one type of virus to choose from (lentivirus, retrovirus, AAV, etc.). When you’ve decided ...sequence for degradation. Viral introduction of shRNA plasmids can induce gene-specific silencing temporarily...Viral expression cassettes typically contain a promoter (inducible or constitutive), your gene of interest...and often a selectable marker. This vector can express your construct in target cells transiently or constitutively...

... smaller size of the plasmids. The Zhang lab, on the other hand, uses a 1 plasmid, single infection system...technology.” GeCKO library for genomic screens Using lentivirus as the delivery vehicle, Neville Sanjana, also... thousands of targets at once. That’s what our plasmids allow.” Wang’s library is designed to target a...have previously been stably transfected for Cas9 expression. The Yusa lab has used their library to screen...

...received over 13,000 plasmids requests for COVID-19 related plasmids. These plasmids have reached labs in...provide synthesized SARS-CoV-2 expression plasmids available for many expression systems: bacterial, yeast,...small collection of plasmids for coronavirus research, including a handful of plasmids that have been in...some highlights of the plasmids shared over the last year and how these plasmids have been used to accelerate... pseudotyping lentivirus with the SARS-CoV-2 spike protein. They deposited the plasmids used for this ...spike protein to improve expression and pseudotyping efficiency. This plasmid was later used by another...circulating and plasmids containing these mutations have been deposited at Addgene. Plasmids deposited from...

... shRNA expression ID Plasmid Description PI 10878 pLKO.1 ‐ TRC cloning vector shRNA expression; Recommended...shRNA expression ID Plasmid Description PI 8452 pMKO.1 puro Retroviral vector for shRNA expression Bob ...shRNA libraries Lentiviral packaging and expression plasmids Background RNA interference (RNAi) is an ...delivery of a plasmid. Below, browse our selection of empty vectors for cloning in shRNAs. Plasmids for constitutive...constitutive expression as well as those that allow for conditional (Cre-lox) or inducible (Tet) expression are...vectors for conditional (Cre-lox) shRNA expression ID Plasmid Description PI 11578 pSico Cre addition ...Lentiviral vectors for Tet-regulated shRNA expression ID Plasmid Description Tet on or off PI 11643 pLVCT-tTR-KRAB...

... to co-express them from one promoter. Specificity The ability of AAV to drive gene expression in specific...cell-specific gene expression and viral infection. Cre or Flp dependent switches control gene expression via switches...transgene expression, since you don’t have to wait for the host to replicate the DNA before expression. Now...non-enveloped icosahedral capsid, that can be used to express genes of interest in cell and animal models. AAVs...laboratory experiments ranging from transient gene expression in a particular cell type to CRISPR genome editing...genes, you can coinfect with multiple viruses to express genes or gene sets over 4.7 kb. Co-infection rates.... On the other hand, if you’re interested in expressing multiple small genes, you can design single rAAV...

...transgene expression. Once packaged in AAV, this pre-recombined DNA could lead to transgene expression in recombinase-negative...sufficient transgene expression but limits the likelihood of recombinase-independent expression. Next-generation...Ready-to-Use AAV Find Ready-to-Use Lentivirus Browse Viral Plasmids ...serotype. Additionally, we check viral transgene expression in transduced cells whenever possible. This post... tube is mislabeled or someone grabs the wrong plasmid? What if bacteria snuck into the prep during the... human error, fragments of DNA from the helper plasmids or the cell genome can be packaged inside the ...) are aligned to the reference sequence of the plasmid used to create the viral prep. In a clean viral...

...analyte), and accumulation of ATP activated GFP expression (receptor), then the transducer could be a microscope...genetically encoded, and luckily, we have pre-designed plasmids for these systems! Ion sensors One common biosensor...retrovirus, can either permanently integrate and/or express long term (up to several years!). Biosensors tend...biosensor to specific tissues or cell (sub)types for expression and downstream monitoring. Nervous about making...ready to deliver your sensor, whether you choose lentivirus, gamma retrovirus, or AAV as your vehicle, we...we’ve got you covered with packaging plasmids! References and Resources References  Bhalla N., Jolly ... as stable systems, rather than transiently overexpressing at the time of use. Another great viral perk...

...precisely control where and when certain genes are expressed to investigate their role in a specific behavior...Which viral vector should be used? Modifying gene expression Generally speaking, one may want to achieve either...application.  Genetic manipulation can also reduce expression levels of certain genes. A popular method uses...allows scientists to draw conclusions due to gene expression in a specific brain area, avoiding side effects...targeting the gene is cloned into a viral shRNA expression vector and the vector particles are delivered...delivered into the brain region of interest. AAV or lentivirus?  Both AAV and lentiviral vectors are used for... a tight spatial and temporal control of gene expression manipulation. They are injected using a stereotactic...

...tetraloop and stem-loop 2; 3. The MS2-P65-HSF1 plasmid which expresses the activation helper protein. SAM can ...Konermann, the first generation of CRISPR-Cas9 plasmids were not designed with gene activation in mind...that will enable even more fine-tuning of gene expression levels and on software for analyzing the data...you are equipped and authorized to make and use lentivirus. Also, note that due to the size of the backbone...

...of the new plasmids, antibodies, and viral preps in the repository through our Hot Plasmids articles. .... Today, it's time for another edition of Hot Plasmids!  Here's what you'll find in this post:  New...., 2023 under CC-BY 4.0 License. Find PE6 plasmids here!  Doman, J.L., Pandey, S., et al. (2023).... to target multiple delivery systems, such as lentivirus and MMLV gag, to specific cell types. It can ...therapies and medical applications.  Find DIRECTED plasmids here! Strebinger, D., et al. (2023). Cell type-specific...pulse-chase experiments, and more. Find LOV-Turbo plasmids here! Next, Qin and Cheah et al. described TransitID...LOV-Turbo into TransitID experiments.  Find TransitID plasmids here! These two developments open up new possibilities...