We narrowed to 754 results for: Tro

...CRISPR/Cas9 to cells for genome editing, and introducing inducible systems into cells, among many other...Desfarges & Ciuffi, 2012). One family of viruses, retroviridae, require integration for their life cycle to... during production (read our blog post on the retroviral life cycle for a more detailed description). ...concerning in clinical settings, as evidenced by retroviral gene therapy trials for X-linked severe combined...immunodeficiency virus type 1 integrase mutants retain in vitro integrase activity yet fail to integrate viral DNA...lessons from provirus and non-integrated DNA. Retrovirology, 1, 13. https://doi.org/10.1186/1742-4690-1-...resources on the Addgene blog Viral Vectors 101: The Retroviral Life Cycle Quick Guide to All Things Lentivirus...

...Stbl4   Electrocompetent cells for cloning and storage of unstable DNA and lentiviral/retroviral vectors...Allows uptake of large plasmids hee   High electroporation efficiency supE44 (glnV44)   Suppression ...cloning and storage, blue/white screening, leucine auxotroph. F- endA1 recA1 galE15 galK16 nupG rpsL ΔlacX74...For cloning into and storage of lentiviral and retroviral vectors or cloning or repeated sequences with...

...Engineering Using CRISPR/Cas Systems has expanded out of control. To further consolidate these FAQs, Le Cong worked...gRNAs with the Cas9 nickase which will together introduce cleavage of the target site with efficiency similar...on each side flanking the change you'd like to introduce. We use Ultramer oligo from IDT (non-PAGE purified...although this might still be okay if you are introducing a selection marker gene. The homology arms should...ideally less than 10 bp away if possible. For introducing small mutations (<50 bp) or a single-point mutation...around 100–150 bp total homology. The mutation is introduced in the middle, giving 50–75 bp homology arms....double digestion. As a successful insertion will destroy the BbsI sites, a double digest should discriminate...

...system along with precisely truncated attP sites to control recombination of the gene construct. A predictable...gene. Effector genes, any gene that you want to control the expression of using the SPARC system. The effector...rest of the SPARC components and expression is controlled by the upstream recombination outcome. Find ...Find SPARC plasmids at Addgene To more precisely control the amount of labeling, you can alter the attP site...effector position, as well as to optogenetically control the depolarization of R2 neurons using CsChrimson...genome engineering in flies Learn more about controlling gene expression in flies Resources on Addgene.org...

...scientists and plasmid lovers - serves as an introduction to plasmids. Plasmids 101 will provide you with...plasmids contain an origin of replication (which controls the host range and copy number of the plasmid)...the lab are usually artificial and designed to introduce foreign DNA into another cell. Minimally, lab-... that scientists can purify and study it in a controlled setting. Read more: Plasmids 101: Protein Tags...environment Produce enzymes that will make specific, controlled changes to an organism’s genome (genome engineering...guide also contains multiple protocols and troubleshooting tips to make plasmid usage as simple and straightforward...

...mimic aspects of organ morphology and physiology in vitro and have emerged as a powerful model for understanding...for doing gene targeting, reporter generation, controllable gene knockouts, and inducible gene inactivation...design. B) Strategy for leak-free transcriptional control. C) Iterative transduction workflow overview. Image...SL-insensitive fluorophore (LSSmOrange) as an internal control and a SL-sensitive fluorophore (circularly permuted...the cpGFP without altering that of the internal control (Fig 1). Because this method relies on the fluorescence...-500nM range) and a high specificity for SLs in vitro, and they further demonstrated that when tested ...

... Many thanks to our guest blogger Jon Backstrom. Jon Backstrom, Ph.D is a biochemist in the Vanderbilt...This post was contributed by guest blogger Jon Backstrom, a biochemist in the Vanderbilt Eye Institute ...oligomerization of biotinylated proteins on avidin can be controlled easier than formation of GST dimers. Avi-Tag ... of added biotins per molecule cannot be well-controlled. The lack of site selectivity typically diminishes...relatively low efficiency of biotinylation, at least in vitro. Our solution was to exploit the best of both worlds...

...activity in the brain was measured with implanted electrodes. The Archon1 voltage reporter allows a way to...cre-dependent version at Addgene! Fluorescent protein and control AAV These fluorescent protein expression vectors...interneurons. These vectors can also be used as the control vectors for experiments encoding a fluorescent-... Addgene in the next 6 months, pending quality control. For an estimate on availability date or to be ... and serotype you’d like to be notified about. Controls Plasmid Serotype Name 114472  AAV5, AAV8 ...Plasmid Serotype Name 119036  AAV1  pAAV-hsyn-flex-Voltron-ST Chemogenetics Plasmid Serotype Name ...

...exhibit profound differences in tropism between cell types (astrocytes, microglia, neurons, etc.) and ...can not recombine with each other. When Cre is introduced to the system it binds one pair of lox sites,...your cells.  Some promoters, such as CAG are very strong and may require less viral particles or relatively...or glial fibrillary acidic protein (GFAP) for astrocytes it is important to test it in the brain region... S (2013) Analysis of Transduction Efficiency, Tropism and Axonal Transport of AAV Serotypes 1, 2, 5, ...and 5 Display Differential Efficiency and Cell Tropism after Delivery to Different Regions of the Central...

... inserted GFP. After characterizing iGluSnFR in vitro, showing that it is selectively activated by glutamate...higher expression levels in bacteria and produces stronger fluorescent signals upon sensing glutamate in ...variants differing in kinetics, affinity, and ∆F/F0. Introduce the two variants in three different membrane-targeting...iGluSnFR3 have higher expression levels and produce stronger fluorescent signals upon sensing glutamate using...addition to the original PDGFR TMD, the researchers introduce iGluSnFR3 in two additional anchoring domains ...induction to express soluble protein CAG: Generic strong promoter. Formulated as AAV-DJ for neuronal expression...

...plasmid deposit please sign up here.        A retroviral mouse CRISPR knockout library Article contributed...phenotype, and this depends on the method used to introduce the gRNAs into cells. Most CRISPR pooled libraries...issue, Sarah Teichmann and colleagues created a retroviral CRISPR knockout library using gRNAs designed ...designed by the Kosuke Yusa Lab. This retroviral library was used to transduce T helper type 2 (Th2) cells from...differentiation factors as the screen readout. This retroviral mouse CRISPR pooled library can be used for screening...concentration of ApoB-LPs has been shown to have a strong correlation with cardiovascular disease and indirect...

...CRISPR-Csm: A novel RNA targeting system by: Susanna Stroik The Doudna lab’s recent work characterized a new...heat-inducible lentiviral plasmids here!   Wu Y, et. al. Control of the activity of CAR-T cells within tumours via...chemically stable tool for microscopy by: Susanna Stroik Microscopy techniques have been moving at the speed...microscopy applications such as super resolution, electron, and expansion microscopy. hfYFP is not only brilliantly...denaturing conditions, high temperatures, and other chaotropic conditions better than most FPs. The group also...expansion microscopy as well as correlative light and electron microscopy and observed high fluorescence resilience...

... to neuroscience. In fact, precise light-based control of biological processes is highly useful in other... to be optimized by Tabor’s lab, making light-controlled gene expression in E. coli easier and more robust...engineered to deliver precise, consistent, and controllable pulses of light. Complicated patterns of light...new pathways with sophisticated gene expression controls. Most cell types do not respond to low-intensity...two systems can be coexpressed for sophisticated control of gene expression. An E. coli light-sensitive...from this publication at Addgene. Multichromatic control of gene expression in Escherichia coli. Tabor JJ...

...sequence NGG TTTV PAM site preserved? Usually Destroyed Yes, Cpf1 cleaves 5' of the protospacer Multiplexing...Cassettes transfection gRNAs expressed from a polycistronic transcript  Requires co-transfection of Cas9...Cassettes transfection gRNAs expressed from a polycistronic transcript Requires co-transfection of Cas9  ...showing that Cpf1 can cleave an array of 4 crRNAs in vitro and when expressed in 293 cells. This allows for...a GFP tag. Four weeks post-infection, there was strong expression of Cpf1 and GFP in the targeted brain...effective method with multiple applications (in vitro, in vivo, transfection, viral transduction) for ...

...need to ensure you have a bead-only control alongside your input control. Elute the DNA after washing and..., which uses formaldehyde or UV light to create strong but reversible bonds. This will keep the protein...of histone modifying proteins, which bind quite strongly to DNA, crosslinking may not be necessary. This...Enzymatic digestion, while gentler on the sample, introduces bias due to the selective nature of the cutting...Antibodies 101: Polyclonal Antibodies Antibodies 101: Introduction to Antibodies Resources on Addgene.org ...

...Be prepared to troubleshoot and be sure to include proper positive and negative controls as you get your...someone to turn to for advice if you need to troubleshoot. What do you know about the vendor and the vendor...anti-tag antibody; and immunoprecipitation-mass spectrometry strategies that isolate an antibody’s immune...complexes and confirm interacting partners by mass spectrometry (Uhlen, 2016). Be sure that the antibody you...The Basics of Western blottingAntibodies 101: Introduction to AntibodiesAntibodies 101: Epitope availability...

...Scientific Support Team here at Addgene, so troubleshooting molecular biology experiments was in the job...core part of our academic training.  Developing strong problem solving skills is just a matter of applying...think back to the last experiment you had to troubleshoot; you probably did not just automatically come...As such, scientists are uniquely prepared to be strong problem solvers both in and out of the lab. The...position. Of course, there are limits to how far strong problem solving skills alone can take you - just...effectively does not mean I am qualified to be an astronaut. But, knowing that this skillset is a part of ...

...regeneration studies, and cancer biology. I was first introduced to bioelectricity when I attended the Developmental...living cells. Since bioelectrical signals are controlled by the opening and closing of ion channel proteins...genetically encoded, light-gated ion translocators to control tumorigenesis." Oncotarget 7.15 (2016): 19575. ...signaling networks: exploiting voltage gradients for control of growth and form." Annual review of biomedical... neural and gap junction protein-mediated cues control polarity during planarian regeneration." Developmental...Vaibhav P., et al. "Transmembrane voltage potential controls embryonic eye patterning in Xenopus laevis." Development...

...research that was done and what it means. The introduction should give you just enough information to allow... addressed. In primary research papers, the introductions should be fairly focused. Consult a review article... etc)? Did they perform all of the appropriate controls? Refer back to the methods section for finer points...automate the process as well. Sign up for eTOC (electronic table of contents) alerts from journals you read...useful papers. For many, this collection is purely electronic. Gone are the days of file cabinets filled to...She is a fan of photosynthesis and writes about autotrophs and biochemistry on her blog. Follow her on Twitter...

...activity in HEK293 by reducing editing from ~30% in control samples to 0-10% in Acr samples. Acr proteins can...When added to human cells six hours after the introduction of Cas9 RNPs, AcrIIA4 reduced off-target editing...specific site. 2. Temporal, spatial, or conditional control of CRISPR activity Regulating Acr proteins by inducible...molecules could achieve rapid, dynamic, and spatial control of CRISPR-Cas activity. For example, fusing AcrA4II...LOV2 domain allows anti-CRISPR activity to be controlled by blue light (Bubeck et al., 2018). 3. Reduce...Your Next Experiment Read more about Optogenetic Control of CRISPR   Resources on Addgene.org Find CRISPR...