We narrowed to 1,017 results for: PLE

...endogenously expressed SCF complex members, an E3 ubiquitin ligase complex. Finally, when auxin is added...fusion protein with the endogenous CRBN E3 ligase complex which will polyubiquitinate the fusion protein....target protein-of-interest and the CRBN E3 ligase complex leading to ubiquitination and proteasomal degradation...acid (IAA), the TIR1 F-box associated E3 ligase complex will ubiquitinate the AID-tagged protein which ...  Chung HK, Jacobs CL, Huo Y, Yang J, Krumm SA, Plemper RK, Tsien RY, Lin MZ. Tunable and reversible drug...Inducible, reversible system for the rapid and complete degradation of proteins in mammalian cells. Proc...

...single-stranded and self-complementary adeno-associated viral vector genome titers by droplet digital PCR. Human... helper viruses like adenovirus (Ad) or herpes simplex virus to facilitate efficient replication. The ..., and the two helper plasmids, commonly called “triple plasmid transfection” (Figure 1).    Figure 1:...quantitative PCR (qPCR) (Aurnhammer et al., 2012), droplet digital PCR (ddPCR) (Lock et al., 2014), or determination...known titer on every qPCR run. Note that the rAAV sample has to be digested with DNase before undergoing... Multiple Authors...

...never enjoyed grant-writing. The more I talked to people with industry experience, the more I heard that...decision, the transition felt scary. While I knew two people from graduate school who were at AstraZeneca and...industry for any length of time, you’ll know more people in industry, which means you’re more likely to ...decade of work. The number of steps, decisions and people involved is staggering. The process is complicated...individual could not bring a drug to the market, so people in industry have to work together. Even as a postdoc...postdoc, you’ll have to work with other people in your group, and they are unlikely to hire you if they... academia, promotions in industry are based on people’s ability to manage others and work as a team as...

...library from Addgene, they’ll need to amplify the sample for further downstream applications. As individual...make the library more difficult to work with. For example, a library in a lentiviral backbone might be more... as individual plasmids in your library. As an example, if you want 100x coverage of your library, and...transformation efficiency (Figure 2).    Figure 2: Example of a serial dilution to estimate transformation...libraries that are similar in size and composition as examples. Table 1 provides an overview of pooled library...protocols you can use as a reference. Table 1: Examples of Different Pooled Libraries at Addgene.  ...Libraries (PDF, 401 KB) Screening Zhang Lab - Multiplexed Overexpression of Regulatory Factors (MORF) Library...

...millions of new cancer cases arise and millions of people die from cancer-related complications each year...cancer, really? While cancer is extraordinarily complex at the molecular level, at the cellular level there...cancer research has focused on picking apart the complex cellular pathways involved in the development of...pathway. Its major appeal is that it enables the simple construction of vectors to study the function of...transcript most commonly found across all TCGA tumor samples is represented, making the results of any such ...biggest challenges facing researchers today is the complexity of the pathways involved in the development of...continue to to build and develop this new resource! Please leave us your thoughts in the comments section ...

...REPAIR and RESCUE can function in a multiplexed manner to edit multiple RNAs. It’s clear that CRISPR RNA ...research. RNA editing advantages RNA editing has multiple advantages over more traditional DNA editing systems...This change specifies the edit to be made when multiple As are present in the gRNA spacer, as ADAR will...target window, possibly due to longer stretches of duplexed RNA. Using an ADAR2DD catalytic mutant, they showed...By mutating residues in ADAR2 that interact with duplex RNA, they hoped to destabilize ADAR-RNA binding...As discussed above, the ability to edit RNA has multiple advantages, including reversibility and use in...

...years ago. Teaching by example No examination of the state of science would be complete without the inclusion...attempts to answer it in five paragraphs, with examples. If that sounds ambitious, that’s because it is...successful scientists have outgoing personalities to supplement their intellectual prowess and scientific skills...Four, “New Science,” Zimmer tackles more recent examples of scientific advances, and posits the way science... much less familiar with as a biologist--as an example of “big science,” a massive, expensive project ...certainly chapters that could warrant their own books. Multiple books have been written about CRISPR alone, and...

...targeting a specific genomic locus, which can require multiple cloning steps (Fujita et al., 2013). CRISPR-mediated...engineered DNA-binding molecule such as the CRISPR complex is expressed in cells to be analyzed, the Fujii...another "in vitro" form. In this form, the CRISPR complex, consisting of a recombinant dCas9 protein and ...interest with dCas9 and a targeting gRNA. One such example by the Xu lab is CRISPR affinity purification in...purification of a specific genomic region is another example of the many tools based on targeting of a genomic...genomic locus through a specific gRNA-dCas9 complex. When combined with next-generation sequencing, this ...research conducted in the Hodaka Fujii Laboratory, please also see our interview with Dr. Fujii. References...

...mediate cleavage rather than the multi-subunit complex employed by class 1 systems. A putative new class...Cpf1 (FnCpf1), they employed an E. coli plasmid depletion assay to discover FnCpf1’s PAM sequence requirements...than following it. The Cpf1 crRNA is also much simpler in structure than Cas9’s; only a short stem-loop...CRISPR systems based on Cas9 were thought to be the simplest CRISPR systems and the easiest to adapt to genome...vectors. In 2017, Zetsche et al. developed a Cpf1 multiplexing approach using a single crRNA array to express...varies widely with gRNA sequence. Both groups used multiple methods to examine Cpf1 off-target editing. First...publication at Addgene. 2. Zetsche, Bernd, et al. "Multiplex Gene Editing by CRISPR–Cpf1 Using a Single crRNA...

... relevant bases within the editing window, for example all C’s will edit to T’s. Thus, it’s important ...knocking out a particular gene isn’t feasible. For example, temporary repression or knockdown allow you to... target genes using dCas9-based activators The simplest of the dCas9 activators consists of dCas9 fused...SunTag system which facilitates recruitment of multiple activation domains to the same genetic locus via... proteins. The Synergistic Activation Mediator complex, generally the strongest activator, consists of...Edit”. Is Cas9 not your Cas of choice?  There are plenty of other enzymes to choose from within the Cas ... Multiple Authors...

.... The optimized plasmid curing method was much simpler and took only ~32 hours instead of ~60 hours. ...strategy for plasmid curing, providing users with a complete editing process. The MUCICAT system While the ...including concatenations into crRNA arrays targeting multiple different sites in E. coli. pCutamp cures the ...Ptet, and Para. The pDonor plasmid pRE57-Ter with multiple terminators can serve to interrupt genes (or gene... pQCascade) with 4 rounds of transformation to complete the N-acetylglucosamine strain library construction...dosages to facilitate swift optimization of strains. Multiple induction systems to choose from. Ready for metabolic...mainly on the development and application of a multiplex genome editing system based on CRISPR-associated...

...used to simultaneously target multiple genes, enabling efficient multiplexed genome editing in a single ...compact genome editors to date. We have developed multiple vector versions for TnpB-mediated genome editing...also operates with a single RuvC-like domain. In simple terms, Cas9 cuts DNA using two "scissors" (one ...a 20-nucleotide (nt) guide sequence that is complementary to the DNA target. The 20-nt guide region binds...Oligo 2: Add 'ggcc' to the 5′ end of your reverse complement guide (see Figure 1). Perform standard phosphorylation...promising for delivery via viral vectors and for multiplexed gene editing applications. As research into TnpB...

...of CRBNmidi alone (A) and in a ternary complex with the multiple myeloma drug mezigdomide and ubiquitination...Construction of solid matrix-antibody-antigen complexes containing simian immunodeficiency virus p27 using...on its own, with drug ligands, and in ternary complexes with other proteins, establishing CRBNmidi as ...mutations.  Here, the researchers were able to target multiple genes in M. abscessus either simultaneously or... (also cyan) embedded in the Titin I10 protein (purple) only when Titin I10 unfolds under tension. Upon... Multiple Authors...

...blood sample, in the best-case scenario [7]. In the clinical setting where patient blood samples are limited...especially if used to make clinical grade hiPSCs. For example, the CytoTune™-iPS 2.1 Sendai Reprogramming Kit...persistence of residual viral material [6]. The complete removal of the residual viral particles and establishment... in the presence of these two factors are not completely understood, the addition of an shRNA for P53 ...grade hiPSCs from easily accessible patient blood samples. All pCXLE vectors listed below in Table 1 are ...cells from CD34+ cells across blood drawn from multiple donors with non-integrating episomal vectors. ...

...the progress of the project. After that rather unpleasant experience I thought: I could do this job so ...companies produce different products and therefore need people with different areas of expertise in technical .... In addition, it can be advantageous to speak multiple languages - that makes communication with customers...are different names for technical support roles. Examples are Technical Service Representative, Technical...optimize” a particular standard lab protocol. For example, someone may attempt super quick incubation times...product in question. This can solve the problem in a simple way even if you don’t exactly know why. Personal... use these communication skills by working with people from many different backgrounds and realizing that...

...surface) of the leaf. This is a simple and easy technique to implement. Success might vary between experimenters...tobacco, Agrobacterium infiltration into leaves is a simple and efficient way to overexpress a gene and evaluate...your experiment (de Ruijter et al., 2003). For example, we did our initial transformations with the GUS... on transformation success. Some reporter gene examples are below. The GUS gene (β-glucuronidase) with...temporal reporters to measure promoter activity, for example. Useful positive controls could include: pSB96 ...expression from Agrobacterium, but not the plant. For example, we have seen strong expression of the GUS gene...

...chalk talk? On the surface, a chalk talk is quite simple. It's you, a dry erase board, some markers (chalk...question, not a vague goal or a "what" question. For example, "we want a better understanding of the Z pathway...acceptable. Aims should be mechanistic, doable by 1-3 people in 1-5 years, and contribute to resolving the overarching...scientists outside your field, and interesting. For example, “there is a significant need for healthy livers...explain why it’s significant and innovative. For example, "while it is possible to grow liver stem cells...chalk talk Perfect your story and delivery. Design simple figures and diagrams to illustrate your vision....also helpful to practice at least a few times with people that are outside your lab and outside your field...

...whole-genome/transcriptome” studies, in which, for example, the activity of all genes in response to an experimental...the globe (Nüsslein-Volhard et al., 2012). One example (which I will come back to) is the one-eyed pinhead...morpholinos. When injected into zygotes, morpholinos complementary to a target mRNA inhibit its translation either...genome is useful for a variety of applications. For example, specific tissues such as blood cells or neurons...regulatory systems have also been successfully implemented in zebrafish. Developmental biology insights ...study profiled 38,731 cells from 694 embryos over multiple stages and reconstructed the patterns of gene ...well-suited for neuroscience investigations. For example, neuronal activity can be tracked in whole brains...

...designated this fragment as the ccdB region (for “coupled cell division”). The encoded toxin-antitoxin system...subunit of DNA gyrase and trapping it in the cleaved complex. This results in DNA breakage and cell death. Conversely... so that the marker can be reused to introduce multiple genetic modifications in the same strain. MazF...translation. Here, Mok and Hok translation is coupled. Translation from the Shine-Dalgarno (SD) sequence...often lost without plasmid addiction systems. For example, Cara Morin and James Kaper used the hok/sok system...Teru, and Sota Hiraga. "Mini-F plasmid genes that couple host cell division to plasmid proliferation." Proceedings...K. Burley. "Crystal structure of the MazE/MazF complex: molecular bases of antidote-toxin recognition....

...maintain itself within host cells.  The plasmid has multiple ori that are functionally differentiated from ...proteins from bacterial hosts. In a plasmid with multiple oris, one may function in one type of host and... the others are functional in other hosts. For example, the pJD4 plasmid harbors distinct origins (ori1... load (Meyer 2009). Figure 1: Idealized examples of a narrow host range plasmid with only one ori... ori and a broad host range plasmid with multiple oris that vary in DnaA-box and iteron structure, host-independent...Acinetobacter calcoaceticus, Actinobacillus pleuropneumoniae, Actinomyces naeslundii, A. viscosus, Aerobacter... hosts in which proteins can be expressed. For example, scientists can incorporate interchangeable antibiotic...