... for something to brighten our day, whether it’s memories of summer or just a bright new plasmid. One ...identified from a high-throughput screen, selected for increased voltage response, brightness, and photostability...sensor dLight1.2 and voltage sensor Ace2N and additional GCaMP6 constructs: pAAV-CAG-dLight1.2 (AAV1)...One area with lots of bright new plasmids is our Fluorescent Biosensors Collection — scientists are always...iGluSnFR3 (Aggarwal et al. 2022). This new version has higher signal to noise ratio and is better localized for...variants optimized for different applications (v82 has higher affinity for glutamate but slower kinetics; v857... post. Figure 2: iGluSnFR3 displays enhanced brightness, expression, and sensitivity compared with earlier...

...relies on ultrasound waves as opposed to light to produce higher resolution images in vivo. “One of the ... result, ultrasound enables higher resolution images at depths up to eight millimeters – not single cells...New York recognized their limitations for peering right through living animals to see their organs – a liver...iRFP713 and iRFP720, is that they absorb and emit light in the near-infrared portion of the electromagnetic... melanin pigments, which absorb the majority of light below 650 nanometers (nm), and the absorbance of...toolbox In the last year, Verkhusha added two far-red light photoactivatable (PA) near-infrared fluorescent ...their fluorescence upon illumination with far-red light. As the researchers described in Nature Communications...

...measured. Archon1 is  bright, exhibits good localization, has high signal:noise ratio, high sensitivity, responds...blue light (445nm). The discovery and applications of protein partners that bind to only the light- or ...Every few months we highlight a subset of the new plasmids in the repository through our hot plasmids...colleagues turned to optogenetics. Through the use of light-sensitive protein domains, they have developed a...system to control CRISPR-Cas9 activity using blue light (Bubeck et al., 2018). They accomplished this by...inhibits CRISPR activity.   With the addition of blue light, the terminal helices of the LOV2 domain unfold,... variant of AcrIIA4.” CASANOVA responds to blue light quickly. Within minutes of inactivating the CASANOVA...

...unprecedentedly-high brightness, bfloGFPa1, and the other with extremely-dim brightness, bfloGFPc1. The...Koike-Yusa et al., Nat Biotech. 2013. Dec 23.   Highly bright and extremely dim GFPs The availability of the...to low-intensity blue light. This approach relies on the incorporation of light-oxygen-voltage (LOV) sensing...made available through Addgene and we want to highlight it for use in your research. Martínez-García... the three dimensional structure of GFP, its brighter mutant S65T-GFP, and the subsequent biochemical ...variety of GFPs with diversity in fluorescence brightness, intensity, excitation and emission spectra. ... MitoTimer reporter using a Tet-on system (pTRE-Tight-MitoTimer) to show its usefulness in cell culture...

...visible light scatter The optical system has both visible and fluorescent light sources. Visible light illuminates...addition to visible light, flow cytometers can have a variety of fluorescent light sources. When interrogated... number based on intensity. Higher intensity events are assigned to higher channels. In a typical assay...experiment. Single-stained samples must be as bright or brighter than any experimental sample. In addition...interrogated with various visible and fluorescent light sources that allow users to assess a cell’s size...and provides useful information about the cell. Light that continues in the same direction as it was initially...size of the cell; larger cells produce more FSC. Light that scatters sideways to the path that it was initially...

...external light activation, you can’t use fluorescence to monitor processes directly affected by light. Long-term...nano-lantern emitted yellow-green light and exhibited 3-5 fold increased brightness over previous BRET probes,...Long-term light exposure can also lead to cellular phototoxicity, and experimental success can be affected...FRET, but it eliminates the need for an excitation light source. Instead, a luciferase substrate like coelenterazine...autooxidize and produce background fluorescence, so higher concentrations of the compound can be added to ... (eNL) described in Suzuki et al. improve the brightness of previous Nano-lanterns by 2-4 fold and permit...fluorescent imaging! These eNL constructs use the brightest characterized luciferase, NanoLuc, and its substrate...

...diagnosis. IDT, 2014. Dwight Z; Palais R.; Wittwer CT. uMELT: prediction of high-resolution melting curves...proficiency with common applications - now let’s dive right in! You know qPCR is most commonly used to measure...statistical significance (Taylor, 2017). With the right conditions and parameters, some people find it can...on hairpin design of probes/primers often offer higher allelic specificity, due to the need for the target...work better for rapid-cycle PCR, such as the LightCycler, as it reduces the primer/probe actions to a ...particular has been reported to work well in the Lightcycler while being relatively easy to multiplex (Thellwell...Agarose gel or uMelt analysis dsDNA dye (Sybr)  High No No No Melt curve Hybridization Probes...

...Arch (green-light activated proton pump) and ChR2 (blue-light activated cation channel) light-activated ...near-complete desensitization in continuous light. ArcLight Q239 - A GEVI based on the fusion of the voltage...deactivated using yellow light (590nm). BLINK2 potassium channel - An optimized blue-light induced synthetic ... is activated by cyan light and exhibits an inactivation maximum of 590 nm light. Aurora and Phobos optogenetic...non-neural cells, ion-based communication happens at a slightly slower scale.  All living cells with a membrane...model (Chernet et al., 2016). There are many other light-based genetic tools for studying bioelectricity ...fluorescent protein. This GEVI features increased brightness and improved plasma membrane specificity. Stable...

...COVID-driven virtual drain. Audience needs high speed internet access. High probability of real-time technical...social media… how do you choose? Which way is the right way?  First of all, I want to put to rest a common... areas of any given country have equal access to high-speed internet. Before you begin any work, make ...communication that you enjoy and that plays at least slightly to your strengths. Otherwise, you’re going to ...Resource Requirements Pros Cons Video Medium to high Uses visuals, audios, and text. Incredibly immersive... writing can difficult.  Photography Medium (high for microscopy) Visually compelling. Easy to convey...to make things more accessible by including, highlighting, or omitting details. Digital or print distribution...

...by an external light source, they have lower background signal and can be used in light-sensitive systems...Every few months, we highlight some of the new plasmids, antibodies, and viral preps in the repository...collection grows By Ashley Waldron We are excited to highlight the Anti-V5 tag antibody produced by our partners...constant domains, the plasmids for the heavy chain and light chain are also available from Addgene!   ...constructs to tag the GOLG-4 protein, mScarlet3 was 50% brighter than the original worm-optimized wrmScarlet. mScarlet-I3...to reduced background signal when tested in more highly autofluorescent tissues like the intestine. ...Expression of fluorophore-tagged GOLG-4 through eight stages of embryonic development. Representative ...

...investigating any biological process. High quality genome sequence A high-quality sequence assembly of the ... Developmental biology insights using lineage tracing in zebrafish To highlight some of the features that...Lineage tracing studies highlight how these advantages can translate into insights in developmental biology...as a useful vertebrate model based on their straightforward husbandry and ability to produce abundant, ... a few of the available genetic resources and highlight two recent breakthroughs in developmental biology...lineage tracing approaches that led to important insights in developmental biology. Microscopy-based lineage... This microscopy-based lineage tracing defined a high-resolution map of early embryogenesis in time and...

...500nm) Light 0.1 Crystal Violet Pre and Post Visible Light 100 Methylene Blue Post Visible Light 100 ... and Post Blue or UV Light 0.5 GelRed Pre and Post (Post recommended) UV Light 0.1 GelGreen Pre and...you to visualize your DNA using UV light – please note that UV light can damage DNA and that proper personal...are having trouble getting colonies, you might want to use higher competency cells. Additionally, if your...bacteria; add chemicals to precipitate out the high molecular weight genomic DNA; filter the remaining plasmid...100 Ethidium Bromide Pre and Post UV Light 0.5 For more information on these stains see the Bitesize...final product is going to be very large (>10kb) you might want to use electro-competent cells instead of the...

...editing rates as high as 83% with NHEJ and 40-50% with HDR. Coselection efficiency is so high that some applications...Every quarter we highlight a subset of the new plasmids in the repository through our hot plasmids articles...27798609 Listen to our mCyRFP1 Podcast Segment New bright monomeric red fluorescent protein - mScarlet! The... new RFP variants for increased fluorescence, brightness, and proper maturation. Three new RFPs were isolated...toolbox Increasingly popular Optogenetics tools use light to alter cell physiology and molecular processes...protein is irreversibly cleaved when exposed to light. Robert Campbell’s lab illustrates the utility of...this caging technique to beautifully demonstrate light-dependent Gal4 transcriptional activation and Cre...

...from an mRNA that forms a pseudoknot. At high copy numbers, high levels of the lncRNA bind to RepZ mRNA ... termination. These types of plasmids tend to be high copy number. Here, we will consider the replication... ori called DnaA boxes. These boxes come in both high- and low-affinity versions, but binding of the low-affinity...: it bends the DNA to redistribute the DnaA from high-affinity boxes to low-affinity boxes, allowing lower... redistributes DnaA (red) from binding primarily high-affinity DnaA boxes (dark red) to also binding low-affinity...DnaA negatively regulates its own expression, so high levels of DnaA act as a feedback mechanism to reduce...stringently controlled plasmids, replication is tightly coupled to the bacterial host's cell cycle, maintaining...

... more carefully controlled. Regulated promoters might act only in certain tissues or at certain times ...them on or off at will with a chemical, heat, or light. In the cell, promoters themselves are controlled...of other promoter elements, and tend to be more tightly regulated. Common promoters for eukaryotes and ...Repressed by ethanol Full length version is strong with high expression. Truncated promoters are constitutive...promoter from maize ubiquitin gene Constitutive  Gives high expression in plants. H1 small RNA expression ...polymerase III RNA promoter Constitutive  May have slightly lower expression than U6. May have better expression...depending on its orientation to your gene. T7lac High levels of gene expression Promoter from T7 bacteriophage...

...species like C. elegans and Drosophila. Light inducible promoters Light is another way to activate gene expression...biology use light to regulate transcription. Red flame plasmid pDawn contains the blue-light sensing protein...switched from an OFF to an ON state, and how you might use these in your research. When you're done with...inducible promoters Chemical agents, temperature, and light are all examples of factors that can lead to the...protein YFI. When light is absent, YFI phosphorylates FixJ, which binds to the FixK2 promoter to induce... preventing expression of a reporter gene. When light is present, YFI is inactive, preventing repressor...transcription to take place. Addgene also has yellow flame light-regulated two component systems designed by the ...

... inducing the brightest fluorescence (Filonov et al., 2014). Broccoli is twice as bright as Spinach2 when...Mango fluoresces nearly as brightly as Spinach, but binds with much higher affinity to its cognate small... their high binding affinity to TO1-biotin. Two of these variants exhibit increased brightness compared...V., et al. "RNA mango aptamer-fluorophore: a bright, high-affinity complex for RNA labeling and tracking...biologist’s kitchen lab - and scientists using aptamers might know this best, as most fluorescent aptamers are...are 10-100 base nucleic acid oligos that bind with high affinity to small molecules to induce their fluorescence... the first aptamer identified to bind DFHBI, is highly unstable in vivo due to poor folding and thermal...

... variant has slightly different activation/inactivation kinetics. For example, the light-gated cation ...exists in several states: closed/light receptive, open, closed/light insensitive. If you deliver a continuous...to “reset” and become receptive to light once again. Therefore, light must be delivered in pulses. An example...the optogenetics coin is materials science-based. Light is delivered to your opsin through a small piece... optic cable implanted into the animal’s skull (right). The fiber optic cable is threaded through—and ...then connected from laser to ferrule to deliver light pulses to the target brain region. Thankfully, due...stimulation (the z-axis bregma coordinate). Because the light that is emitted from the end of the fiber optic ...

...the blue light molecule much, much, much more light-sensitive. Then you can use dim blue light and red ...the discovery of a light-sensitive opsin from algae that is sensitive to red light instead of blue, which...to tinker with brain circuits using the power of light. In a recent article published in Nature Methods...electrical activity of neurons on or off using pulses of light. By activating a neuron or set of neurons, you can...that all of these molecules are sensitive to blue light. Many have tried to make green molecules, but then...red light together. Understanding the brain through optogenetics research Addgene: What are some of the ... deep part of the brain, making it sensitive to light. When they illuminated this region, the animals ...

... not only brilliantly bright, but it also withstands denaturing conditions, high temperatures, and other...microscopy as well as correlative light and electron microscopy and observed high fluorescence resilience to ...retains folding and fluorescence in high molarity GdnHCl, at high temperatures, and in the presence of...Every few months we highlight a subset of the new plasmids, antibodies, and viral preps in the repository...CRISPR-Csm doesn’t degrade proximal RNAs nor does it have high levels of off-target effects, both of happen with...of antibodies we produced at Addgene and, as you might suspect, it has been one of our most commonly requested...8R] as another option for targeting GFP. But why might you need a second GFP antibody?  Similarities:  ...