...Multiple Multi-luciferase reporter vector including transcriptional reporters for NF-kb, TGF-b, c-Myc, ...them useful as transcriptional reporters. However, they are not secreted, so cells must be lysed or the...decay reporter and control plasmid utilizing Firefly luciferase. Transcriptional Element Screening Optimized...plasmids : Genome-wide screening of enhancer activity using a Firefly luciferase reporter. These plasmids improve... element or gene of interest into to create a luciferase reporter. Unless otherwise noted, these plasmids...transcriptional regulation and bioluminescent reporters. Plasmid...Luciferase Plasmid Collection You may also like... Fluorescent Proteins Blog: Luciferase 101 Blog: Technologies...

...RFP ratio: a decrease in GFP fluorescence relative to RFP fluorescence suggests an increase in autophagic...flux, while an increase in GFP fluorescence relative to RFP fluorescence suggests a decrease in autophagic...autophagy. The development of several fluorescent autophagy reporters now allows for the measurement of autophagic...to lysosomal proteases is key to its pH-dependent fluorescence. In neutral environments, dimeric Keima...Rosado, Carlos, et al. "Rosella: a fluorescent pH-biosensor for reporting vacuolar turnover of cytosol and...mammalian cells using a single fluorescent biosensor." Scientific reports 5 (2015): 12397. PubMed PMID:...photoconversion Light sheet fluorescence microscopy Resources on Addgene.org Fluorescent protein plasmids and...

...fundamentally the reporter construct monitors the expression of a nearby gene. The reporter gene serves as...any way. For example, if green fluorescent protein (GFP) was the reporter gene and green cells were observed...gene. The actual reporter construct can be something phenotypic (such as a fluorescent protein or something...introduce reporters throughout an organism’s genome. These reporters have historically been used extensively...stimulate the expression of the reporter. The system is made up of a reporter gene flanked by a minimal promoter...isolated and screened. Depending on the marker, traps can be used as high-throughput screens to introduce...promoter – all rely on the random integration of a reporter construct in close proximity to an endogenous ...

...NLuc to fluorescent proteins to make better reporters and the second couples NLuc to fluorescent proteins...better biosensors. NanoLuc®-fluorescent protein fusions for imaging Fluorescent proteins are fantastic imaging...more penetrative orange fluorescent protein fusion (OgNLuc). Both fluorescent proteins can be excited ...has dubbed these fusions LumiFluor Reporters. The LumiFluor reporter constructs were designed to be packaged...for infection of host cells. The fluorescent moiety of the BRET reporter was used to sort successfully infected...cyan, yellow, green, orange and red fluorescence. The paper reports a wide variety of applications of the...transfer of energy from one fluorescent protein to a compatible fluorescent protein. FRET can be measured...

...available. Detection methods Many calcium reporters use a single fluorescent protein (FP), requiring only a simple.... This sensor switches from green fluorescence to a red fluorescent state only by simultaneous (1) user-controlled...expression, and compatible with constitutive or Cre-dependent expression. These sensors have also been re-...Genetically encoded calcium indicators (GECIs) are fluorescent proteins linked to calcium binding proteins (... (e.g., calmodulin or troponin C). Their fluorescent readouts are used to monitor calcium concentrations...GCaMP6 family of sensors are commonly used calcium reporters with different kinetic parameters. There are three... of the sensor, and (3) subsequent change in fluorescence only occurs at high calcium concentrations. ...

...CRISPRa depended on continued expression of dCas9 and gRNAs whose expression varies depending on transgene...construct and the fluorescently-labeled gRNA multiplex. Read the paper in Scientific Reports Find the plasmids...The new editors reported in this study have ~10-100-fold lower average Cas9-independent off-target DNA ...DNA editing, ~5–50-fold lower levels of Cas9-dependent off-target DNA editing, and maintain ~50–90% of average...stem cells Lindy Barrett’s lab developed new fluorescently labeled piggyBac vectors to deliver uniform ... lines with AAVS1-integrated, inducible, and fluorescent dCas9-KRAB and dCas9-VPR for quantifying and ...cost-effective screening methods, David Liu’s lab identified and validated CBEs with reduced Cas9-independent deamination...

...artifacts. If you're using green fluorescent protein or Enhanced Green Fluorescent Protein (GFP/EGFP) for a transcriptional...strategies can help you make better fluorescent fusion protein reporters for your studies and greatly lower... second FP, often decreasing the fluorescent signal from the first FP and increasing the signal from the... transcriptional reporter or as a general cytoplasmic label of cells, there's no problem. You're OK. However... means that your POI fused to GFP or another fluorescent protein (FP) could be forming dimers in cells... a particular type of molecule to form a dimer depends on its molecular affinity, termed the dissociation... Figure 1: Each problem with dimerizing fluorescent proteins is illustrated. 1) Membrane proteins...

...mouse strain in which Cre expression causes the induction of both DsRed fluorescence and GFP shRNA. Mice...level. Upon Cre/loxP recombination, each transgene expresses one of three possible fluorescent proteins,...This system uses two plasmids in tandem, with Cre-dependent inversion determining between 0 and 2 FPs expressed...Cas9 gene editing. Dr. Masahito Ikawa has created a GFP reporter plasmid for scientists to validate the ... recombines independently, generating tens or hundreds of possible combinations (depending on the number...mutagenesis libraries or to insert heterologous genes, reporters, or other markers into a target genome. They did...engineer a wide variety of GFPs with diversity in fluorescence brightness, intensity, excitation and emission...

...split mCherry fluorescent reporter, enabling the detection of phosphorylation-dependent protein-peptide...phosphosite, mCherry fluorescence is restored, so interactions can be discovered by fluorescence-activated cell...central position. Introducing Hi-P for screening phosphorylation-dependent protein-protein interactions. The...proteome-wide identification of phosphorylation-dependent interactions. Barber KW, Muir P, Szeligowski RV...NEDD4-2; 111705-8 ) is available from Addgene. Screening kinases. The mode #1 phosphosite library can also...

... versions of this reporter to target the K9 (pcDNA3-K9 histone methylation reporter) and K27 (pcDNA3-K27...The K9 reporter uses the HP1 chromodomain to recognise lysine 9 methylation, while the K27 reporter uses...and Alice Y. Ting. "Genetically encoded fluorescent reporters of histone methylation in living cells."...) that brings the fluorescent units of the FRET pair together and causes an increase in the FRET signal... inactivated, but different phenotypes result depending upon whether the improperly imprinted genes come...techniques include chromatin immunoprecipitation, fluorescent in-situ hybridization, methylation-sensitive ... methylation in living cells. Essentially this reporter construct consists of one substrate (the histone-derived...

... microbes rely heavily on fluorescent protein genes. These optical reporters have limited deep-tissue ... consists of a library of reporter plasmids, each containing two fluorescent proteins and four repeats...collection of reporter plasmids is amenable to Golden Gate assembly for turning the single-input reporters into...field further with the creation of the largest miRNA sensor collection reported to date, containing all...Neuron. 2019. PubMed PMID: 30704911. Acoustic reporter genes for noninvasive imaging of microbes in mammalian...contributed by Lukas Morgan  Listen to the acoustic reporter gene podcast segment In recent years, scientists...organism, the expression of some genes can change depending on the location. Current methods for visualizing...

...Campus reminds us that, although fluorescent proteins are incredibly useful, chemical fluorophores can... a Monomer? The Top Three Ways Your Favorite Fluorescent Protein Oligomerizes in Cells GFP oligomerization...Recombinase-based State Machines Enable Order-dependent Logic in vivo Nathaniel Roquet from the Lu Lab...Lab at MIT developed plasmid based systems that report the order in which specific chemical inputs are ...recombinase-based state machines could potentially be used to report upon and/or engineer “temporally ordered biochemical...derived from Zika virus and, upon binding, produce reporter proteins. These toehold switches can be used to...viral replication and transcription as well as reporter genes. To learn how you can use minigenomes in...

...along with a reporter, such as a fluorescence gene or a resistance gene. These reporters allow you to ...stream of the IRES or 2A element (depending on the placement of the reporter gene). Addgene's collection offers...at equivalent levels upstream of an IRES fluorescent reporter for easy detection.5  Just released: Addgene's... using IRES elements. Further, scientists have reported lower expression of the downstream cistron due...eukaryotic cells, and, although some scientists report close to 100% cleavage with some of these peptides...the choice of specific 2A peptide will ultimately depend on a number of factors such as cell type or experimental...co-express your gene of interest along with a fluorescent protein or selectable marker, it is easiest to...

...Cre-dependent. If Cre-dependent vectors are used in transgenic animals that express Cre in only specific cell types...neurons with an AAV9 vector expressing a Cre-dependent fluorescent marker. Injection into a conditional mutant...product or are fluorescent. While dyes usually spread unidirectionally, there are some reports of bi-directionality...expression. Fluorescent proteins are widely used for tracing and their expression is often made Cre-dependent...4107258. Lo, L., and Anderson, D.J. (2011). A Cre-dependent, anterograde transsynaptic viral tracer for ...biotinylated dextran amines (Veenman et al., 1992) or fluorescent molecules like rhodamine-isothiocyanate (RITC...germ agglutinin WGA, Schwab et al., 1978) or fluorescent molecules such as Fluoro-Gold (Schmued and Fallon...

...approach to selecting a bright fluorescent protein and a photostable fluorescent protein. In the third post.... In corals, the red fluorescent proteins form tetramers. In general, fluorescent proteins have a natural... whorls is probably concentration dependent and cell-type dependent. Therefore, it is good practice to...established true monomeric fluorescent proteins. Concluding remarks Many fluorescent proteins are available...your research. But be aware that the fluorescent proteins reported in publications are only characterized... Addgene Blog Which Fluorescent Protein Should I Use? Choosing Your Fluorescent Proteins for Multicolor... with Fluorescent Tags Find Plasmids for Subcellular Localization Browse the entire Fluorescent Protein...

...El-Samad lab has deposited plasmids encoding fluorescent reporters and other proteins controlled by GAL4 and...estradiol and progesterone and showed that reporter expression is dependent only upon its cognate inducer. Since...random variation, or noise, in reporter expression. To do so, one places reporter expression under the control...expression of a reporter protein, it is possible to change the amount of noise in reporter gene expression...interest led to rapid, targeted degradation that is dependent on proteasome function. TRIM-away compared favorably...deliver the target antibody. TRIM-Away reagents and reporters are available from Addgene - we hope they will...in HeLa and MEF cells. For further reading on fluorescent microscopy techniques, the first author of this...

...localization peptides, and reporters that drive the expression of the fluorescent protein by a specific transcription...labeling Increased neuronal expression using Cre-mediated AAV Targeted Evolution method (M-CREATE) dCas9...-4.   Increased neuronal expression using Cre-mediated AAV Targeted Evolution method (M-CREATE) by: Brian...multiplexed version of the Cre-mediated AAV Targeted Evolution method (M-CREATE), in which one applies both...proximity labeling   By Ashley Waldron Proximity-dependent labeling has become a powerful approach for proteomic... a cleavage free manner. The dCas9-SSAP system reports an efficiency of up to 20%, comparable to commonly... For a one kilobase insertion, the researchers reported an editing error rate of less than 0.3%, both ...

...evaluation of shRNA efficacy using fluorescence; use with pGFPns-reporter for cDNA target rtTA On Gu 60495...the TRE, permitting transcription. Tetracycline-dependent promoters are developed by placing a TRE upstream...system. Tetracycline off is also known as the tTA-dependent system. Tetracycline On System In 1995, Gossen...residues of tetR were important for tetracycline-dependent repression. Mutating these residues led to the...on system is also known as the rtTA-dependent system. Experimental Tips Choosing a tet system If your gene...system are: Plasmid containing a tetracycline-dependent promoter upstream of your gene of interest tTA...system are: Plasmid containing a tetracycline-dependent promoter upstream of your gene of interest rtTA...

...mentioned above (Sanger sequencing, qPCR, and fluorescent reporter-based) each come with their advantages and...opinion, the most convenient strategy is the fluorescent reporter based method since it lends itself to cheap...that uniquely express selectable and/or sortable reporters in certain states of interest. For example, in...a unique combination of red, blue, and green fluorescent proteins in each state. The three methods of...detection with flow cytometry. However, expressing reporters will consume cellular resources and therefore ... chromosome backbone) contains the register. Depending on the desired application, the inducible promoters...expected? The successful implementation of GRSMs depends on the reliability of the parts used to build them...

...Chemigenetic sensors are better classified by their fluorescent reporter and the mechanism of their voltage sensing...Cellular communication and function depend on potential energy. Cellular activities like action potentials...and the control of voltage gated ion channels all depend on transmitting and sensing electrical potential...neuronal pathways and the function of certain cells depends on the ability to monitor their signals – electricity... elements: a voltage-sensing component and a fluorescent output. The GEVI family of gadgets includes Ace...VSD-based sensors consist of a VSD fused to a single fluorescent protein (FP). This class of GEVI was the earliest...of red-shifted indicators, and brightness of fluorescence all have room for improvement. These sensors...