...reversibly degrade proteins of interest fused to a degron, a domain that induces degradation. In plants, ...ligase complex, promoting its interaction with the degron sequence and ultimately its degradation. While ... needed for AID. By tagging proteins with a 7kD degron called mAID and expressing the essential plant ...channels that can be used to silence the activity of neurons. Earlier this year, the Hegemann lab and collaborators... cysteine.  With the introduction of MerMAIDs, neurons can be turned off quickly and with high precision... a valuable optogenetics feature when studying neuronal activity. The researchers identified MerMAIDS ...

...there was strong expression of Cpf1 and GFP in the targeted brain region and ~75% of neurons were co-transduced...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  ...multiplex edit 3-4 genes in a primary culture of mouse neurons and in vivo. For these experiments, cells were ...-transduced with Cpf1 and GFP.  Of these neurons, ~15% had indels at all 3 targeted loci.  See the chart...

...Murphy’s law definitely applies: what can go wrong, will go wrong. The trash removal service slams dumpsters...useful transferable skill. Being able to stand up in front of an audience and convey your ideas clearly and...project. So whenever you freeze, or when things go wrong, ask yourself, “What next step will help me achieve...term “diving board” so you end up acting it out in front of a very confused group of visiting Chinese undergrads...incorrectly on slides 8-16. Your notes disappear. The wrong version of your slide deck gets uploaded. Your mic...

...Tet-on transgene vector. Degron tags Tagging relevant proteins with a degron tag can provide inducible...removed. Degron tag systems include small molecule assisted shut off (SMASh), auxin-inducible degron (AID)...SMASh degron tagging vector! Destabilizing domains Destabilizing domains (DD) are akin to degron tags,...

... RNA-sequencing to identify neuronal class and deep sequencing of neurons from each class to fully characterize...Addgene.  C. elegans open resources The C. elegans Neuronal Gene Expression Map & Network (CeNGEN) The CeNGEN...comprehensive gene expression atlas of every C.elegans  neuron. Scientists are currently doing this using two ...this system a modified TIR-1 protein can degrade degron-tagged proteins in an auxin dependent manner. This...Gene Expression Map of an Entire Nervous System." Neuron 99.3 (2018): 430-433. PubMed PMID: 30092212. PubMed...

...across the membrane of cardiac cells. Similarly, neuronal signaling is carried out by action potentials ...perceived signals to physical actions. Understanding neuronal pathways and the function of certain cells depends...be directed to specific types of cells, such as neuron subtypes, thanks to viral serotype specificity ...available VSDs has been successful even at the single neuron level in vitro. In vivo monitoring has also been...membrane as well. Another flavor of these tools is Voltron, which uses rhodopsin as the voltage sensing component...voltage is particularly physiologically relevant in neurons, myocytes, and endocrine cells. It follows that...

...norepinephrine (NE) expressing neurons and GFP (green) to label neurons transduced by each serotype. ...challenge to scientists. What works well in one cell or neuron type may not work well for a neighboring cell type...express ChR2(H134R) in A2 norepinephrine expressing neurons. We tested four serotypes containing an expression...84%) were both excellent at infecting our target neurons (Figure 1). However, this one set of experiments...Perspectives on Data Sharing in Deep Brain Stimulation. Frontiers in Human Neuroscience, 14(December), 1–12. https...

...Chemical Synapses in the Brain. Neuron 68:843–856. https://doi.org/10.1016/j.neuron.2010.11.021  Gambarotto D...techniques like super-resolution microscopy or electron microscopy. Since its introduction in 2015 by ...2010). How do you overcome this resolution limit? Electron Microscopy (EM) is one option, but the equipment...helped researchers gain a deeper understanding of neuronal synapses, centrosomes, parasites, and more (Bertiaux... Ashley Waldron...

...biological studies, including investigations into environmental pollutants and health, embryo growth, brain ...example, specific tissues such as blood cells or neurons can be fluorescently labeled, allowing researchers... for neuroscience investigations. For example, neuronal activity can be tracked in whole brains of swimming...high-resolution mapping of the brain by uniquely labeling neurons. Zebrafish are also amenable to manipulation of...consequences of activating or silencing specific neurons in living larvae. In addition, zebrafish have been...PubMed PMID: 18845710. Kim, Dal Hyung, et al. "Pan-neuronal calcium imaging with cellular resolution in freely...

...make them ideal for studying in a laboratory environment. We’ve covered five model organisms in a previous...candidate for introductory teaching and research environments (Idalia & Bernardo, 2017).  Escherichia... disease-relevant genes in such a controlled environment displays how essential the frog is as a biomedical...genome, and is easily maintained in a laboratory environment. What sets Arabidopsis apart from other plant...Regeneration and Aging Research: A Mini-Review. Gerontology, 57(6), 565–571. https://doi.org/10.1159/000323761...the Axolotl Model for Regeneration and Aging. Gerontology, 66(3), 212–222. https://doi.org/10.1159/000504294...

...Access to Projection Neurons. Neuron, 92(2), 372–382. https://doi.org/10.1016/j.neuron.2016.09.021 Walters..., 2023). It may be a good choice for targeting neurons, with high transduction frequencies in the central...to target the central nervous system, including neurons, AAV5 is generally the best choice for targeting... is derived from AAV2 and engineered to infect neurons and undergo retrograde transport, traveling up ...

...molecule at all, leading to inconclusive results, erroneous conclusions, and wasted time. In fact, many have...recommendations for improving antibody use and reliability (Roncador, et al., 2015; Taussig, et al., 2018; Uhlen, et...that antibody works as expected, you will see a strong antibody signal in the retinal sample, but little... your antibody (B). Your results show a lovely, strong band from the cells transfected with your protein...Nature 518:27–9. https://doi.org/10.1038/518027a  Roncador G, Engel P, Maestre L, Anderson AP, Cordell JL... Ashley Waldron...

...mouse neurons. D) Membrane voltage traces of action potentials in a HcKCR1-expressing mouse neuron without...optogenetic silencer in experiments with mouse cortical neurons, far out-performing previous attempts to engineer...of HcKCR1-EYFP and tdTomato expressed layer 2/3 neurons in mouse. C) Action potential frequencies with ...antibody anti-c-Myc [9E10] now available! by: Ashley Waldron The majority of Addgene's more than 50 antibodies...

...at Addgene, I get to see everything that can go wrong with a plasmid. One of the most frequent problems...smaller plasmids are formed. Viral vectors are prone to recombination In lentiviral vectors, recombination... elements. Knowing how to amplify recombination-prone plasmids and testing your prep can save you the ...H (2012) Remarkable stability of an instability-prone lentiviral vector plasmid in Escherichia coli Stbl3...Genetic recombination of bacterial plasmid DNA: electron microscopic analysis of in vitro intramolecular...

... Frank Graham showed HEKs display many immature neuron-like qualities, which is inconsistent with kidney...is next to the kidney and is known to have some neuronal qualities.) While HEKs probably aren’t a great...researchers using the cell line. What the HEK is wrong with it?   All cultured human cell lines are susceptible..., et al., Preferential transformation of human neuronal cells by human adenoviruses and the origin of ...Produce Recombinant Proteins and Viral Vectors. Front Bioeng Biotechnol. (2021) (9):796991. DOI: 10.3389...

...equipment always helps. But how does one build a strong collaborative research team motivated by similar...your lab team and foster an effective working environment? Tom Ellis was offered a start-up package that...ways to improve the lab. “An effective working environment for research is totally dependent on communication...while others are more productive in a handsoff environment. “Treating people fairly does not mean equally...selectively picking the first couple of hires to build a strong, collaborative team. And take advantage of the ...

...error prone PolI3M that’s fused to a nicking Cas9 (nCas9). Similar to mutagenesis with error prone PCR,...bacterial growth conditions. Expression of an error prone polymerase I (PolI3M) is capable of mutagenesis ...PCR, EvolvR uses an error prone polymerase to introduce mutations. However, by using nCas9 to direct PolI3m...   Figure 1: EvolvR consists of an error-prone polymerase (PolI) that is fused to a nicking Cas9...

...signaling pathways in specific cell types (typically neurons). While all of these viruses are popular, pAAV-...pAAV-hSyn-DIO-hM4D(Gi)-mCherry can be used to induce neuronal silencing in vitro and in vivo (Krashes et al..... The human synapsin promoter further specifies neuron-specific expression of the DREADD (Kügler et al...Lowell BB. Rapid, reversible activation of AgRP neurons drives feeding behavior in mice. J Clin Invest..... Human synapsin 1 gene promoter confers highly neuron-specific long-term transgene expression from an...

...plasmid for expression in forebrain GABA-ergic interneurons (83895). See our Serotype Testing AAV inventory...with ever more efficient tools for light mediated neuronal control. The advantages of optogenetics lie in...system that allows for specific manipulation of neurons. We have expanded our AAV collection to include...targets expression of ChR2 to forebrain GAB-ergic interneurons through the mDlx enhancer element. Plasmid...DREADD acts through the Gq pathway and promotes neuronal firing. Plasmid Serotype Name Depositor 44361...

...skills in strong relationship-building and influence. You’ll need a large deal of influence and strong relationship-building...software product management was either. Coming from a strong background of undergraduate research straight to...energized from interacting with others, and have strong communication skills to tailor your message to ...implementation review. The developers discuss which front-end and back-end coding frameworks to use. I mostly...