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  1. Advanced Uses of Cre-lox and Flp-FRT - A Neuroscientist’s View

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    ...Visualizing single neurons in the living brain   Figure 1: Plasmid mix to label neuronal morphology (...labeling of cortical neurons and their synapses.   How can you label a single neuron and its synapses...DNA’, is derived from the yeast S. cerevisiae (Gronostajski and Sadowski 1985). The FLP system works in ...neuroscience Manipulation of specific cell types in neuronal networks Double inverted vectors expressing light...manipulate specific cell types (populations) within neuronal networks in living animals. To accomplish such...electroporation. To achieve sparse labeling of individual neurons one could reduce the plasmids amounts to reduce...efficiency. However, if the cellular structure of a neuron and its synapse should be visualized at the same...
  2. Hot Plasmids Spring 2024

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    ...www.janelia.org/open-science/voltron-and-positron.  Both Ace2N-mNeon-A122D and Voltron2 are available as Cre-dependent...33714753. Voltron2 voltage sensors By Brian O’Neill New viral vector preps expressing the Voltron2 voltage...performance.      Figure 4: Structure of Voltron or Voltron2 conjugated to a JaneliaFluor dye emitting...HaloTag domain.  Find Voltron2 plasmids here! Abdelfattah, A.S., et al. (2023) Neuron, 111(10), 1547–1563... SARS-CoV-2 research New viral vector preps of Voltron2 voltage sensors AAV vectors for targeting microglia... to facilitate SARS-CoV-2 research   By Ashley Waldron Ready-to-use recombinant antibodies were recently..., are now available! Building on their earlier Voltron indicators (a fusion of Ace2N and HaloTag, Figure...
  3. Important Considerations in Optogenetics Behavioral Experiments

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    ... Inhibition of neuronal activation is a little more complicated since basal neuronal activity in your...advent of optogenetics, ICSS can now be done with neuronal cell-type and temporal precision. In ICSS, a test...actually stimulating or inhibiting the targeted neuronal population prior to behavioral testing. One method...a reliable and widely used molecular marker of neuronal activation and is a sufficient readout to proceed...readout may be necessary to prove inhibition of neuronal activation, but it may still be possible to use...example, as shown in this study [3], hippocampal neurons are activated when the mouse is placed in a novel... the authors were able to show that HR blocked neuronal activation in the novel context as determined ...
  4. 15 Hot Plasmids from 2017

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    ...linked to disease, including motor neurons (-3mnx1/Hb9), pan neuronal (elavl3/HuC), microglial/macrophage...specific neuronal subtypes was not available until now. In order to target specific neuronal subtypes ...type SOX2 with either a SOX2-VP16 (strong activator) or SOX2-HP1 (strong repressor) fusion protein, the authors...interest. In order to improve the study of specific neuronal cell types involved in neurodegenerative diseases...), and astrocytic (gfap).This toolbox adds new neuronal tools to the expanding gateway compatible vectors...to express the transcription factors from a polycistronic cassette. However, it was unknown whether variations...variations in gene expression from these polycistronic cassettes could lead to comparable experimental...
  5. Hot Plasmids May 2018 - Optogenetics, Decaffeination, Biosensors, and Fluorescent Protein Tools

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    ...expression in E. coli. This operon is under the transcriptional control of a strong constitutive promoter (...powerful tool used by neuroscientists to interrogate neuronal networks. A major drawback of this method is the... the stray excitation of neighboring neurons in opsin dense regions. To overcome this issue, Shemesh et... C-terminus of the protein. Unlike the diffuse neuronal expression observed with wild type CoChR, soCoChR...describe a novel system for the precise mapping of neuronal networks at the single-cell level. Various soCoChR...PMID: 29184208.        Synthetic decaffeination operon Article contributed by Angela Kaczmarczyk  Listen...Listen to the synthetic decaffeination operon podcast segment With the help of engineered microbes, used...
  6. Lighting Up Cell Signaling with Photoswitchable Kinases

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    ...cyan light, Dronpa dimers dissociate, Dronpa goes dark, and MEK1 is active or “on.” Dronpa’s color thus...Additionally, dimeric Dronpa is less prone to aggregation in cells compared to the tetrameric Dronpa it was derived... by attaching two dimerizing photoswitchable Dronpa (dron= a ninja term for vanishing, and pa= photoactivation... the Dronpa dimers interact and lock MEK1 in an “off” setting by blocking the active site. Dronpa also...similarity. For these three kinases, photoswitchable Dronpa domains were inserted at sites homologous to the...here. In addition, it could be possible to use pdDronpa to control other kinases and proteins. Zhou et...proteins could potentially be created by inserting pdDronpa domains into loops of a protein of interest. If...
  7. Multicolor Animals: Using Fluorescent Proteins to Understand Single Cell Behavior

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    ...2011) labels Drosophila neurons at the neuroblast stage, meaning that neurons of the same lineage will...cell types, and research applications. Labeling neurons with Brainbow The Lichtman group developed the ...iteration of this technology in 2007. Used to label neurons in the cerebellum of mice, the team appropriately...2007). Brainbow uses a Cre/lox system to label neurons. Genes encoding the fluorescent proteins are organized...the truncated YFP. In Thy1-Brainbow mice, some neurons expressed as many as 16 copies of the injected ...entire length of the cell, even axons of up to 100 microns in length. Some challenges of the Brainbow system...cells, the expression level of these XFP is not strong enough for all applications. To resolve this, similar...
  8. 22 Hot Plasmid Technologies from 2014

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    ...’s lab has enabled progress on both fronts – as a genetic electron microscopy (EM) tag providing superior...presynaptic region of one neuron and postsynaptic region of a different neuron, come into proximity in ...Druckmann et al., Neuron. 2014. Feb 5;81(3):629-40.  Next-Gen Brainbow toolkit for neuronal imaging Joshua...Howarth. Shining a light on channelrhodopsins – Chronos & Chrimson Through de novo sequencing of 127 algal...These tools include a pair of channelrhodopsins, Chronos and Chrimson, which have enabled the group to perform...perform independent two-color optical excitation of neurons. The paper describes five previously unknown ...and Scherffelia dubia (SdChR). ShChR, nicknamed Chronos, is blue and green light-excitable and has faster...
  9. Plasmids 101: The Promoter Region – Let's Go!

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    ...Additional considerations CMV General expression mRNA Strong mammalian expression promoter from the human cytomegalovirus...some cell types. EF1a General expression mRNA Strong mammalian expression from human elongation factor...promoter hybrids. CAG General expression mRNA Strong hybrid mammalian promoter Constitutive  Contains...activate promoter. Ac5 General expression mRNA Strong insect promoter from Drosophila Actin 5c gene Constitutive...Drosophila. Polyhedrin General expression mRNA  Strong insect promoter from baculovirus Constitutive  ...calmodulin-dependent protein kinase II promoter Specific Used for neuronal/CNS expression. Modulated by calcium and calmodulin...mammalian EF1a promoter. GDS General expression mRNA  Strong yeast expression promoter from glyceraldehyde 3...
  10. Cancer and the Immune System: Deciphering the Relationship

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    ...effector cells in the tumor microenvironment. Tumor cells in this environment produce growth factor signals...inflammatory cytokines - interleukin (IL)-12 and interferon gamma (IFNγ) - and eliminate cancer cells. 2....responses that eliminate early stage cancer cells, chronic inflammatory responses can augment tumor growth...development are observed in autoimmune disorders. Chronic inflammation due to autoimmune disorders is also...Strategic immune evasion mechanisms - The tumor microenvironment Through interactions with the immune system...and thrive within what is known as the tumor microenvironment - a vital atmosphere created by the cancer...recruit regulatory T cells (T-regs) into their microenvironment by producing C-C motif chemokine 22 (CCL22...
  11. Getting Started with Behavioral Neuroscience: Standardizing Experiments and Using Viral Vectors

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    ...AAV1 and AAV9 serotypes have strong neuronal tropism, AAV5 targets both neurons and astrocytes, and AAV4 ...activation of neurons in a specific brain region. One option is to overexpress a gene that alters neuronal function...introduced in neurons and depending on the choice of channel protein, they can trigger neuronal excitation...certain tests and is not always recommended.  Environmental influences like bright light or noise should...(vendor or in-house mouse breeding colony) Environment Olfaction, temperature, humidity, noise level...factors. Another option uses optogentics to control neuronal activity, commonly via light-activated ion channels...broad cell tropism and has a high percentage of neuronal targeting after stereotaxic injection. Find more...
  12. Viral Vectors 101: Chemogenetics

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    ...example, lowers the threshold for neuronal signaling and causes neuronal burst firing. Other DREADDs, such...DREADDs for Neuroscientists. Neuron 89:683–694 . https://doi.org/10.1016/j.neuron.2016.01.040 Saloman, J. ...method for manipulating cell activity, especially neuronal firing. What is chemogenetics? Chemogenetics ...decreased heart rate (Redfern et al., 1999) or altered neuron excitability (Zhu et al., 2014). The system allows...Studies looking at immediate effects of specific neuron firing are better equipped using fast-acting optogenetic...tissues, cell types, or even subcellular regions of a neuron need to be targeted. Targeting is also important...common target for chemogenetic manipulation are neurons where chemogenetic receptors can be used to alter...
  13. New and Upcoming Viral Vectors - Spring 2019

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    ... of densely labeled neurons because background fluorescence from inactive neurons is reduced. 105322... of densely-labeled neurons because background fluorescence from inactive neurons is reduced.  Additional...calcium sensors and can be used for imaging of small neuronal processes like dendrites and axons. Find the 7b...Archon1 voltage reporter allows a way to monitor neuronal electrical activity through imaging. Since it’...mDlx enhancer restricts expression to GABAergic interneurons. These vectors can also be used as the control...Plasmid Serotype Name 119036  AAV1  pAAV-hsyn-flex-Voltron-ST Chemogenetics Plasmid Serotype Name ...flex PSAM4 GlyR IRES EGFP Chimeric channels for neuronal manipulation 119742  AAV5  AAV SYN PSAM4 GlyR...
  14. New and Upcoming Viral Vectors - September 2019

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    ... of densely-labeled neurons because background fluorescence from inactive neurons is reduced. This variant...50457-AAV5): The Synapsin promoter directs broad, neuronal expression. AAV pCAG-FLEX-EGFP-WPRE (51502-AAV5...element directs expression in forebrain GABAergic interneurons. pOTTC1032 - pAAV EF1a Nuc-flox(mCherry)-EGFP...AAV5 and 112677-AAVrg): The EF1a promoter directs strong, ubiquitous expression. This is a Cre-dependent...(jGCaMP7) continue to be useful for monitoring neuronal activity in the form of calcium dynamics. (For...fluorescence and can be used for imaging small neuronal processes (dendrites and axons). jGCaMP7c exhibits...method for understanding how this molecule impacts neuronal circuits. We recently packaged a constitutive ...
  15. A Primer on Optogenetics: Introduction and Opsin Delivery

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    ...VTA GABA neurons disrupts reward consumption. Neuron. 2012;73(6):1184-94. doi: 10.1016/j.neuron.2012.02.016..., for example, the firing of a specific type of neuron in a specific brain region can result in a particular...wherein mouse initiated optogenetic activation of a neuronal circuit either does or does not result in a reward...consider whether you wish to stimulate or inhibit neuronal firing, the cell-type specificity, the kinetics...suggests that Arch may more effectively silence neurons than HR. Delivering your opsin  The standard opsin...manipulations to help restrict expression to a specific neuronal subtype. However, the length of promoter that ...construct is limited to only a few kb so attaining neuronal specificity through promoter-directed expression...
  16. Viral Vectors 101: Calcium Sensors

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    ...to monitor? Neurons, for example, have unusually fast calcium dynamics, whereas non-neuronal systems tend...populations of neurons in vivo to study circuit activity. CaMPARI is available for neuron-specific expression...and target a defined population of cells (e.g., a neuron subtype) and specific subcellular compartments ... rapid measurements, like action potentials in neurons. jGCaMP7 and jGCaMP8 are also available with improved...improved version named CaMPARI2 is available for neuron-specific expression in both zebrafish and mammalian...studies. There are different versions optimized for neuron-specific, astrocyte-specific (GCaMP6 only) or mammalian... to measure changes in calcium specific to the neuronal cell body or the axon (GCaMP6 only). The genetic...
  17. Viral Vectors 101: AAV Variables That Matter

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    ...great in a GABA neuron, for instance, may not transduce the adjacent norepinephrine neuron at all. There... for Defense Behaviors. Neuron, 93(1), 33–47. https://doi.org/10.1016/j.neuron.2016.11.045 More resources...or inducible.  Ubiquitous promoters  These often-strong promoters offer the most flexibility for expression...Promoters such as chicken-beta-actin or CAG can drive strong early gene expression, but can become methylated...expression can still occur (Fig. 3) - for example, some neurons can transiently express TH - so you’ll need to ...expression in both oligodendrocytes (Olig2) and neurons (NeuN). Image from Griffin et al., 2019, Figure...Second, high efficiency transduction coupled with a strong promoter, may result in overexpression of your ...
  18. Avoiding the Dark Side of Fluorescent Protein Fusions with mOX FPs

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    ...whether and how well your FP will function in the environment where it will localize [4]. Most investigators...in or were engineered for use in cytoplasmic environments. However, approximately 40% of human (and most... localize to chemically distinct subcellular environments, including the organelles that compose the secretory...to interfere with the function of resident ER chaperones and may impact cellular function or possibly ...consequences of misfolded dark FP fusions including erroneous quantitation of fusion protein levels. It remains...these compartments using CRISPR systems should strongly consider using moxFPs over standard FPs for the...optimizing fluorescent proteins for subcellular environment. Currently, she is a SPIRE postdoctoral fellow...
  19. New Neuroscience Tool: The iGluSnFR3 Glutamate Sensor

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    ...soluble protein CAG: Generic strong promoter. Formulated as AAV-DJ for neuronal expression. CAG-FLEX: CAG... the ability to monitor glutamate release from neurons and other brain cells in vivo. Glutamate plays ...synaptic communication and can trigger other forms of neuronal signaling and regulation. To create iGluSnFR, ...responsiveness, they showed that iGluSnFR could detect neuronal activity in C. elegans, zebrafish, and mice. ...higher expression levels in bacteria and produces stronger fluorescent signals upon sensing glutamate in ...membrane-targeting flavors – PDGFR, GPI, and SGZ, to improve neuronal membrane trafficking and localization. Improved...iGluSnFR3 have higher expression levels and produce stronger fluorescent signals upon sensing glutamate using...
  20. A Practical Guide to Optimizing AAV DIO and FLEx Vector Expression

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    ...cell types (astrocytes, microglia, neurons, etc.) and even neuronal phenotype in a mixed population (Burger...have experienced that some vectors are much more prone to recombination and can exhibit recombination levels...your cells.  Some promoters, such as CAG are very strong and may require less viral particles or relatively...tyrosine hydroxylase (TH) promoter for catecholamine neurons or glial fibrillary acidic protein (GFAP) for astrocytes...vectors for off-target expression.  Some classes of neurons transiently express TH during development or have...there is promoter activity. Additionally, some neurons express low levels of GFAP, which can lead to off-target...off-target expression of the packaged transgene in neurons and glia.   Check the recombination rate The first...
  21. Bringing Sustainable Practices to the Lab: Innovations

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    ...their environmental impact on a scale of 1-10: the lower the score, the lower the environmental impact...receive the product? How can we consider the environmental impact of those processes? Who even regulates...years since the creation of the United States Environmental Protection Agency, which started making regulations... can choose products that are better for the environment without sacrificing performance or quality. To...destined for a landfill. However, Triumvirate Environmental developed Red2Green, a program that uses this...lumber for things like truck bumpers, plastic adirondack chairs, and synthetic turf. I was truly amazed... a practice which we now realize is a major environmental disaster. We should continue to examine our ...
  22. Top Requested AAV of 2017: pmSyn1-EBFP-CRE

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    ...pouring in! While our lentiviral service is going strong, the AAV service has shown incredible growth this...be used to drive Cre expression specifically in neurons. AAV Delivered Cre - A useful tool for neuroscientists...way. For example, say you would like to label neurons with mCherry and you already have a transgenic ...entirely new mouse line with mCherry controlled by a neuron specific promoter or you could breed your Cre-dependent...line with a separate line that expresses Cre in neurons, but both of these options would be very time consuming...and therefore mCherry will only be expressed in neurons. Indeed, this AAV provides you with a means to ...expression of any Cre-dependent gene of interest in neurons. This ability to be paired with any Cre-controlled...
  23. Fluorescent Protein Travel Awards - FLiPs and Fluorescent Protein Biosensors

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    ... make enzymes controllable by tethering them to Dronpa, a fluorescent protein known to oligomerize when...To do so, she generated fusion proteins in which Dronpa domains flanked either side of an enzyme’s active...ultraviolet light exposure caused oligomerization of the Dronpa domains thereby occluding the enzyme active site...the enzyme. Later exposure to cyan light stopped Dronpa oligomerization and turned the enzymes back on....optimized these first generation FLiPs by mutating Dronpa to lower its aggregation tendency, increase its.... She additionally showed that these enhanced “pdDronpa” domains could be placed in amino acid loops on...iterations of FLiPs only used N- and C-terminal dronpa fusions. With these various designs, Xin has been...
  24. Scientists Map the SARS-CoV-2-Human Interaction Network

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    ...Manon Eckhardt and Melanie Brewer from the QBI Coronavirus Research Group at UCSF. It’s been only a few ...-- all while maintaining social distance.  From front to back: David Gordon, Jeff Guo, Fengbo Zhou, Paddy...scientists from more than 20 labs to form the QBI Coronavirus Research Group (QCRG) to do just this.  In under... York. We want YOU to join the fight against coronavirus We are just at the beginning of understanding...stick together (6 feet apart), to fight the new Coronavirus. To help get you started, here are a few key ...clone selection. Expression is driven by a fairly strong EF1alpha promoter. We have verified expression ...lab's Addgene page to get notified!) Attacking coronavirus, one FedEx package at a time Since the publication...
  25. GPCRs: How Do They Work and How Do We Study Them?

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    ...proteins, work in different neurons and neuroglia, and have different effects on neuron activity (firing or inhibition...in a specific neuronal subtype in the brain that can either activate or inhibit neurons. You can find ...DREADDs for Neuroscientists. Neuron 89:683–694 . https://doi.org/10.1016/j.neuron.2016.01.040 Takeda S, Kadowaki...chemogenetic studies to probe the relationship between neurons and specific behaviors. Chemogenetics uses genetically... small molecules to either activate or inhibit neuronal firing. As GPCRs are the largest class of signal...concise table of DREADDS and their activity in neurons on Addgene’s chemogenetics guide.     References...
  26. 27 Hot Plasmids from 2016

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    ... Jason Moffat at the University of Toronto has created the Toronto KnockOut (TKO) CRISPR Library. This...for neuronal systems. To develop a new single-component tool for magnetogenetic control of neurons, the...sequences outside of their endogenous chromatin environment and relies on the fact that enhancers function...J Genet Genomics. 2016. PubMed PMID: 26887506 Toronto KnockOut (TKO) CRISPR library In order to identify...Natl Acad Sci USA. 2016. PubMed PMID: 26884157 Aronshtam, et al. Nucleic Acids Res. 1996. PubMed PMID: ...but turns more red in lower (e.g. vacuolar) pH environments that inactivate SEP and leave only DsRed to ...the Co-IP compared of a control Co-IP indicates stronger interaction. The system has been demonstrated ...
  27. Hot Plasmids and Viral Preps - March 2021

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    ...inducible degron (AID) system in C. elegans By Alyssa Cecchetelli The auxin inducible degron (AID) system...Tracking cell-cell interactions New auxin inducible degron system for C. elegans New CRISPR plasmids New items...cell types including endothelial cell-pericyte, neuron-astrocyte, and cancer-stromal cell pairs.   ...   Schematic showing the auxin-inducible degron system. Image from Ashley et al., 2021.   The...spatiotemporal expression pattern and an auxin inducible degron (AID) sequence attached to your protein of interest...presence of auxin, this TIR1 complex binds to the degron sequence ultimately leading to the degradation ...the AID* (minimal 44 amino acids auxin inducible degron sequence) tagged proteins. Degradation of your ...
  28. Interview: Ed Boyden on Optogenetics, Neuroscience, and the Future of Neuroengineering

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    ...activity of neurons on or off using pulses of light. By activating a neuron or set of neurons, you can see...activate a set of neurons, maybe we could treat patients suffering from Parkinson's, chronic pain, deafness...independently control the activity of two populations of neurons at once. Addgene spoke to Boyden about this latest...contributes to behavior or pathology, and by disabling neurons you can figure out what they are necessary for....examples where you want to activate two sets of neurons and see how different parts of the brain or the...
  29. R Bodies: Membrane-Rupturing Microscopic Tools

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    ...Much later, it was shown that putting the operon under a strong promoter results in high R-body expression...machines that can extend and retract over many microns in response to pH changes without the need for ... are composed of 4 proteins encoded in a single operon and were first expressed in E. coli in the 1980s... scaffold that changes shape in response to environmental conditions. More speculatively, R bodies might...of armor to protect their host from predation (Petroni et al 2000). With a better understanding of how...new ways to interact mechanically with their environment.   Jessica Polka is a Postdoctoral Research ... Plasmids from This Publication at Addgene 4. Petroni, G., Spring, S., Schleifer, K.-H., Verni, F. & ...
  30. Cancer, Inflammation and Immunity - Harnessing the Body’s Defenses to Fight Cancer

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    ...from a state of chronic hepatitis caused by hypernutrition and alcohol consumption. Chronic inflammation ...cancer and defeat it we need to attack it from all fronts!”  Here I aim to compile a few highlights from ...researchers are preparing and carrying out this multi-pronged attack. Inflammation and tumorigenesis The conference...about alcohol and obesity as driving factors for chronic inflammation which in turn suppresses cancer immunosurveillance...surface receptor PD-1. This immunosuppressive environment in turn may represent a favorable state for tumor...Resources. Studying the immune system and tumor microenvironment Tumor Immunosurveillance and Defense With ...Friedmann-Morvinski about tools used to analyse the tumor microenvironment (one of the plasmids on her poster is pTomo-Ras...
  31. Tagging Optogenetics and Chemogenetics Receptors: Fluorescent Proteins and Other Options

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    ...our observations at the electron microscope showed us that, in monkey neurons, the large majority of the...are powerful tools to modulate the activity of neurons and other brain cells. Since the opsins or chemogenetic... we used AAV viral vectors to express hM4Di in neurons of rhesus monkeys or mice (Galvan et al., 2019)...found a larger proportion on the membrane in mice neurons, most of them were still cytoplasmic. Use of ‘small...hM4Di‐mCherry at the plasma membrane in monkey neurons, we next hypothesized that a different tag fused... of interest the membrane.   Figure 1: Electron microscope images showing ultrastructural localization...protein of interest, an alternative could be bicistronic expression of the fluorescent protein with the...
  32. Important Considerations When Using AAVs

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    ...oligodendrocytes, microglia, astrocytes, neurons, and inhibitory neurons. Your choice of AAV will depend upon... thanks to our guest blogger Katrina Armstrong. Katrina Armstrong is a Neurophysiology Msc Student at ...post was contributed by guest blogger Katrina Armstrong, a Neurophysiology Msc Student at the University... These AAVs may move toward the cell bodies of neurons (retrograde transport) or towards the synapse (...a larger amount of vector using a serotype with strong CNS tropism (4). If axonal transport is not desirable...neuroanatomical tracing (11). With the ability to produce neuron specific transfection using Cre-lox, this new method... method could allow for labeling of neurons showing a very specific phenotype. Previous neural tracing...
  33. 28 Hot Plasmid Technologies from 2015

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    ...plasmids: Luc (intronless; CMV-LUC2CP/ARE and Luc-I (intron-containing; CMV-LUC2CP/intron/ARE). The Luc-I...Luc-I intron is a chimeric β-globin/immunoglobulin intron that has been commonly used in constitutive splicing...Multimerization via the SunTag system also shows strong upregulation of gene activation when fused to dCas9... simplifies the process of creating complex polycistronic lentiviruses. The MuLE toolbox contains a series...induced-pluripotent stem cells (iPSCs) to take on a “neuron-like” phenotype when targeting genes involved in...in mammalian cells, yeast, and Drosophila. Synchronization of secretory protein traffic in mammalian cells...disrupts this interaction and thus triggers a synchronous release of the reporters which can then be tracked...
  34. Save Time with Transient Plant Leaf Transformations

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    ...tobacco. Studying gene function in C. roseus is strongly limited by the availability of methods. Infection...reporter gene examples are below. The GUS gene (β-glucuronidase) with histochemical GUS-staining is a very useful...days in the dark after the transformation. Use introns Plant promoters can be recognized or leaky in Agrobacterium...Agrobacterium, but not the plant. For example, we have seen strong expression of the GUS gene with the commonly used...Agrobacterium. This can be easily avoided by the use of introns, which will be removed from the transcripts in ...plants, check out this interview with the Nicola Patron, who shared the MoClo plant kit via Addgene. For...well working protocol in your lab, it can become a strong tool for your research. Feel free to add further...
  35. Fluorescent Proteins 101: Aptamer Fluorophores

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    ...electrochemical sensors for emerging environmental pollutants.” Front Chem. 2014; 2: 41. PubMed PMID: 25019067...variety of applications, including detection of environmental pollutants, drug discovery, and disease diagnosis...instead of protein-based fluorophores, including: Environments with higher temperatures, since proteins can...Jonathan. “RNA Fluorescence with Light-Up Aptamers.” Front Chem. 2016 Jun 28;4:29. PubMed PMID: 27446908. PubMed...systems using immobilized aptamers.” Biosens Bioelectron. 2011 May 15;26(9):3725-36. PubMed PMID: 21419619.... PubMed Central PMCID: PMC4800036. 12. Wang, Rongsheng E., et al. “Aptamer-Based Fluorescent Biosensors...assay for prion diseases diagnostic”. Procedia Environmental Sciences. Volume 12, Part B, 2012, Pages 1348...
  36. Plasmids 101: Plasmid Incompatibility

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    ...factor RepA binds to the iteron to initiation replication. However, in iteron-containing plasmids,  RepA... regulation system involving antisense RNAs or iterons that inhibits replication when copy number in the...   Figure 1: The replication problem. Iterons are repeated sequences within the origin of replication...del Solar et al., 1998). There are two models for iteron-based plasmid replication control that are not ...the second model, termed the “handcuff” model, iterons of two separate plasmids bind to the same RepA ...these plasmid types? Read our Plasmids 101: Environmental Plasmids blog). Scientists have designed small...replication and transfer systems and host taxonomy. Frontiers in Microbiology 6: . https://doi.org/10.3389/fmicb...
  37. Plasmids 101: Stringent Regulation of Replication

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    ... pSC101 copy number is regulated by iterons. RepA binds to iterons and recruits host proteins to the ori...is initiated at the ori and may or may not be synchronized with the replication of the host cell's chromosomal...RepA binds to directly repeated sequences called iterons located near the ori and directly interacts with...check, high copy numbers can allow RepA to link iterons from two different plasmids, forming a pair of ...either of them (Park et al., 2001). The number of iterons encoded on a given plasmid influences how high ...host, as no additional components are necessary. Iterons are not the only tool to stringently regulate plasmid... numbers, RepA proteins form complexes between iterons on different plasmids in a process called "handcuffing...
  38. Multifunctional Neural Fibers: Modulation from Gut to Brain

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    ...neural fibers developed by the Anikeeva lab are microelectronic fibers made from thermoplastic polymers that...desired thickness. The fibers are drawn with microelectronics included in them for downstream control – ...trended with expression of the opsin in the target neurons, validating the functionality of these inputs and...Success! They also observed reliable optically evoked neuron activity for at least several months after implantation..., P., & Chandrakasan, A. P. (2020). Modular Optoelectronic System for Wireless, Programmable Neuromodulation... Anikeeva, P. (2019). Flexible fiber-based optoelectronics for neural interfaces. Chemical Society Reviews...A., … Anikeeva, P. (2023). Multifunctional microelectronic fibers enable wireless modulation of gut and...
  39. Viral Vectors 101: Optogenetic Tools

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    ...the first light-activated tool for controlling neuronal activity, channelrhodopsin-2 (ChR2), a blue light-activated... exposed to blue light, this channel activates neuronal activity by opening and allowing positively charged...channels (rhodopsins) either excite or inhibit neuronal activity upon exposure to light within a specific...hyperpolarize the cell (Mattis et al., 2012). Bicistronic vectors combine excitation and inhibition by ... when you want to activate or inhibit specific neurons without needing to consider secondary signaling... off-target activity. Whether you are studying neuronal signaling or cancer pathways there are a number...2022). Red Light Optogenetics in Neuroscience. Frontiers in Cellular Neuroscience, 15, 778900. https://...
  40. Visualizing Protein Turnover In Situ

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    ... How do you visualize protein turnover within a neuron? For my PhD I studied a synaptic protein that is... to study how protein turnover is regulated in neurons. Protein turnover is the combined effect of protein... I wanted to measure it’s levels in its normal neuronal context. In other words, I needed a method for...for visualizing protein turnover in situ in neurons. The classic approach for studying protein turnover... whole population of one particular protein in neurons or in any other cell type. However, we cannot distinguish... newly synthesized proteins in rat hippocampal neurons." Nature neuroscience 13.7 (2010): 897-905. PubMed...Robert H. Singer. "Single β-actin mRNA detection in neurons reveals a mechanism for regulating its translatability...
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