... molecule degraders” recruit the proteasome to destroy targeted proteins. We previously developed small...locus-specific knock-in of FKBP12F36V. Detailed protocols describing the use of these plasmids are available here....Nabet et al., Nature Chemical Biology additionally provides detailed information on cloning procedures used...degradation, and lead author on the recent paper describing the dTAG technology. Follow him on twitter @behnamnabet... 29581585. This reference contains the full description of the dTAG system. The dTAG system was employed...Conditional Mouse Models CRISPR 101: Cas9 Nickase Design and Homology Directed Repair Plasmids for endogenous...

...approaches to gender equality over the past several decades: “Fix the Numbers of Women” focuses on increasing...-art methods of sex and gender analysis and 2) provides case studies with concrete examples of how gender...using both sexes in basic research is excellent design), but they used all female stem cells—this was ...environmental factors. How to include sex in experimental design Recognizing the importance of analyzing sex, the... above, another excellent resource for research design is Kalpit Shah et al., “Do you know the sex of ...Corporations need to integrate these insights into product design. But eyes have been opened—and we cannot return...

...monitored over time by sequencing the barcodes in the population (all barcodes can be amplified using the same...with roughly 73 million semi-random, 30 bp DNA barcodes that are integrated into target cells' genomes...ClonTracer system, lentiviruses containing these barcodes infect the target cell population at low multiplicity...replicate populations, they found that similar barcodes consistently made up the majority of the resistant...resistant populations. In fact, 88% of the barcodes within the resistant populations were shared by at least...

...repository that will help epigenetic researchers custom design their own engineering plasmid tools in about a ...cloning, the Haynes lab’s Protocols.io protocol describes an elegant system that allows for the assembly... cells that make up the human body and G-baToN provides versibility in tracking and modulating the interactions... inhibitor response 1 (TIR1) expressed under a desired promoter with a specific spatiotemporal expression...interests via CRISPR techniques. This tool-kit includes an array of FPs as well as additional useful tags...libraries encode target sites flanked by randomized nucleotides in place of a protospacer adjacent motif (PAM...

...of 3’UTRs, the Arizona State University (ASU) Biodesign institute developed a 3’UTR database specifically...specifically for C. elegans. This database provides detailed information about C. elegans 3’UTRs structures, ...imaging processing. The combination of these methods provides unique opportunities to develop computational ...Addgene In addition to the wealth of open resources described above, there were many plasmid tools presented...self-excise, and covalently ligate their flanking peptides together (Wang et al., 2018).” With the split ...continuously curated as we continue to receive plasmids designed for the worm so check back frequently for what...

... Neurodegeneration Research Collection , which includes other areas of this research topic. Michael J....Browse or search the table below. ID Plasmid Description Gene/Insert Industry PI Return to top...

...degradation of the tagged protein. Degrons are destabilizing domains which will be identified by various ...proteasome for degradation. Like several other tags described here, the dTAG uses a small molecule, a tag, and...protein for imaging. Given its popularity and widespread usage, a proteolysis-targeting chimera (PROTAC... The auxin-inducible degron (AID) system is the oldest system reviewed here. The AID system relies on ...Thankfully, this is a hot topic with many resources! In Bondeson et al, they review all the tags discussed here...informed choice! References and Resources References Bondeson, D.P., Mullin-Bernstein, Z., Oliver, S. et al....

...genetic locus when complexed with an appropriately designed small RNA, or guide RNA (gRNA). The gRNA conventionally...exploited to either inactivate a gene or introduce a desired genetic alteration. Listen to Our Podcast Interview... truncated such that it exhibits 15 or fewer nucleotides of complementarity to a target site, Cas9’s DNA...truncated gRNAs Importantly, the use of truncated guides has largely been demonstrated to result in decreased.... An additional benefit of the use of truncated guides is that they can be delivered into systems that...Cas9 species. Within a single cell, full-length guides targeting one set of genetic loci can be introduced...

...amazing project. You’ve written your protocols, designed your experiments, and prepared your reagents. ...following cloning methodologies have been extensively described in other articles; so if you are interested in...homology. You might also consider FastCloning, first described by Chaokun Li and colleagues. Here you simply ...-generated insert. When mixed, topoisomerase I provides both the endonuclease and ligase function while...restriction enzymes. In Golden Gate cloning, the desired end construct does not contain the type IIS recognition...This feature biases the reaction in favor of the desired assembly to make the process close to 100% efficient...

...insertion site, the corresponding gRNA might be undesirable due to potential off-target sites or low predicted..., resulting in lower overall genome frequency. Despite the progress we’ve made, there is still a need ...bacterial polymerase subunit ω. This construct is designed to activate transcription from the accessory plasmid... stringency and promoting survival of only the broadest-targeting variants. For the final round of evolution...spectinomycin resistance. xCas9 3.7 showed the broadest PAM range, recognizing NG, NNG, GAT, and CAA PAMs...scaffold. With an NGG PAM, xCas9 3.7-VPR constructs modestly outperformed SpCas9-VPR in activating transcription...

...Snapp. Stop using EGFP/GFP for fusion proteins! Despite multiple studies in high profile journal articles...world examples of these situations in cells are described below (See Figure 1).   Figure 1: Each...of environmental changes that the biosensor is designed to detect resulting in false positives [2, 4]....Addgene has many of these constructs and they are designated mEGFP, mVenus, etc. A word of caution is needed... not necessarily monomeric, even if they were described as monomeric in a publication. The gold standard... [2]. Some affinity assays used in the papers describing the FPs above including molecular sizing columns...

... to imagine designing 10-gene systems with quantitative precision. There are few designers of biological...made news in March with a publication in Science describing the development of “transcriptors,” transistor-like... and observed that we humans find it useful to describe and interact with the world using words like OR...The future of synthetic biology Addgene: You've described the pieces in these kits as the beginnings of ...laying down a challenge to its users and to the design community: If you can begin to reliably put together...

... crRNAs for Cpf1-mediated cleavage are 42-44 nucleotides in length, about the same size as Cas9’s, but...SpCas9. Since Cpf1 doesn’t require a tracrRNA, crRNA guides are only ~42 nt long. Direct synthesis of these... than that of the ~100 nt crRNA/tracrRNA hybrid guides needed for Cas9 function. Since both Cpf1 and its... endjoining (NHEJ). Cas9-mediated NHEJ usually destroys the PAM site due to its proximity to the cleavage...chance” mechanism might improve the frequency of desired HDR edits, but the possibility has not yet been...to examine Cpf1 off-target editing. First, they designed gRNAs with single and double mismatches throughout...

...available from this depositor's lab. Plasmid Description Gene/Insert Vector Type PI Publication Back to... plasmids from this depositor's lab. Plasmid Description Vector Type Selectable Marker PI Publication ...

...site, and titer. Addgene’s Viral Vector Service Besides being complex to use, viral vectors are also difficult...which include calcium and voltage sensors, are the broadest category of viral vector tools, which may help... for vectors in the “controls” category, which includes things like viral vectors that deliver GFP. ...most requested serotypes (Figure 4), our catalog includes vectors in a variety of serotypes, including the...early engagement with UPenn to our launch of the redesigned Data Hub in 2022. It’s influenced the way we ...viral vector educational resources, from eBooks to guides to protocols, which was why I was so excited to...

...of data. In this two-part blog series, we will describe the importance of bioinformatics at Addgene and...each week. Our internal software was originally designed to help us work in large formats, such as 96-well...Thanks to the flexibility inherent in Nextflow’s design, we were free to use multiple programming languages...alone! The benefits of containers are they are designed to ‘contain,’ in a complete package, all of the... who or which computer ran the process. Another design choice we appreciated about Nextflow was how effortless...small with a print statement and tested that the desired data was available so we felt confident we were...

...their BioRxiv post, the authors generated and described several improved genetically encoded fluorescent...Takayuki Iwaki, Kimiko Hara, and Kazuo Umemura have described a simple method which alleviates the inclusion...contains the signal peptide of MBP, followed by the desired nanobody (NB) sequence, an AviTAG, and 6xHis tag...expression can vary significantly depending on transgene design and delivery. To overcome these variations in human...Addgene The MoClo CRISPR/Cas Toolkit for plants includes 95 plasmids consisting of CRISPR/Cas nucleases... uses CRISPRi to control gene expression. This includes a CRISPRi-based synthetic oscillator, bistable...

...promotes scientific sharing The EDGE Program is designed to expand the number of systems that can be studied...genotype to phenotype studies New organismal systems described at this meeting were extraordinarily diverse. ...generating interesting drug-like compounds especially cardenolides which have been used to treat heart disease....disease. Caterpillars that eat milkweed generate cardenolides that tastes yucky (and is poisonous) for bird ...control who a female wants to mate with and if she decides to raise their offspring or leave them to their...of basic evolution of fungi and eukaryotes and includes the fungus that is killing frogs all over the ...

... that can help with the transition.” The lab-to-desk-job culture shock is a real thing. I experienced...challenging than lab work at times.” Sometimes a desk job is defined as sitting down at a computer and...consider as you make this transition. Of course, a “desk job” could entail any number of career paths--industry...colleague Mike Lemieux told me, “I find that at a desk, it is much easier to stick to timelines. This is...from legal, Diana in finance, and Wu in graphic design. You could dread these interactions, but you could... move away from calling non-academic careers and desk jobs “alternative careers.” Realistically, based...

...iPSCs. The generation of human iPSCs (hiPSCs) now provides an opportunity to develop and use patient-specific...for reprogramming purposes. In this post, I’ll describe an episomal vector system that increases reprogramming...the CytoTune™-iPS 2.1 Sendai Reprogramming Kit designed to generate clinical grade hiPSCs is sold at $11,000...replication, OriP and the EBNA-1 sequence that encodes for the DNA binding protein EBNA-1 [7]. These two...reported efforts. The pCXLE episomal system thus provides a practical way to generate transgene-free, virus-free...factors they encode Episomal Plasmid Addgene ID# Encodes pCXLE-hSK 27078 SOX2 and KLF4 pCXLE-hUL 27080...