...other systems because the system modulates protein post-translationally, unlike many other methods. Secondly... tags has become increasingly popular for the modulation of endogenously and exogenously expressed proteins...systems. Degron tags are one of many tools to manipulate protein expression. These tags offer several ...Secondly, these tags can be used to manipulate endogenously and exogenously expressed proteins. Thirdly...degradation. Since its first use as a protein regulation tool, AID has gone through an additional round...

...any antibody (positive population) or won’t bind anything (negative population). It is strongly recommended...immunologists, allowing for the rapid analysis of cell populations within a heterogenous tissue type, such as PBMCs...moreso than other methods, the identification and isolation of rare cell types. In recent years, full spectrum...for better resolution of positive and negative populations, Careful panel design In both conventional and...create difficulty when trying to resolve those populations. PE/Cy7 and BB515, on the other hand, have a ...

...DNA in your sample relative to your normal control - you can set up your calculations so that your assay... one-copy ‘housekeeping’ gene), and quantified relative to a 'normal' sample with known copy number set...nice thing about qPCR is that since it measures relative quantification - that is, since you’re looking...all double-stranded DNA, while Taqman uses an intercalating dye, with a fluorescent probe and quencher to...provides absolute quantitative data, instead of relative quantification.  Even though no one option is ...

...important to understand that these platforms, or pieces of a platform, that are now available exist within...well-characterized bacterial transcription and translation initiation elements. Open science with BioBricks...replication of DNA, transcription of DNA to make RNA, translation of RNA to make proteins. That's central to everything... transduce otherwise invisible molecules into regulation of gene expression including growth or fluorescence...gene expression via standard transcription and translation initiation." Nature Methods. 10, 354-360 (10 ...

...about how they became grant professionals, mid- to late-career grant professionals have similar stories ...’s requirements as well as state and federal regulations. A grant professional’s job duties can range ...close-out a grant. Grant professionals also build relationships with foundations and government grantmakers....keep up-to-date on the changing guidelines and regulations impacting grant management, we track grant opportunity...amount of grant funds available and the increasing regulatory process for grants has fostered the demand for...worked for a health department and then for the regulatory services department as our healthy homes program...

...However, retroviral vectors can still direct the translation of proteins because they integrate this reverse-transcribed...then use cell machinery for transcription and translation. Here, we will discuss several aspects of the...The genome of viral tools are often not virology related at all, and you will instead get expression of ... Cullen, B. R. (1992). Mechanism of action of regulatory proteins encoded by complex retroviruses. Microbiological... sarcoma virus: purification of the virus and isolation of the nucleic acid. Proceedings of the National...Initiation of synthesis with 70 S viral RNA as template. Journal of Molecular Biology, 79(1), 163–183....

...more focused format, where all students work on malate dehydrogenase (MDH), a citric acid cycle enzyme...recently purified MDH to study the impact of phosphorylation on the enzyme’s structure, function, and interaction...offering excellent training in improvisation and the latter making him a more effective motivator (“be like...conducting NIH- and NSF-funded research on planarian flatworms both in and out of the classroom.   References...characterization of a fusion protein: Histidine tag, malate dehydrogenase, and green fluorescent protein. Biochemistry...laboratory – a semester-long research approach using malate dehydrogenase as a research model. Biochemistry...

...vector - overall, I spent two years testing and reformulating our AAV construct to optimize performance in...Working at Addgene has shown me this was far from an isolated case. Our scientific support team answers hundreds... AAV questions every year on a range of issues related to AAV vectors, many from people trying to answer.... (2020). Methods Matter: Standard Production Platforms for Recombinant AAV Produce Chemically and Functionally...its occurrence during vector purification and formulation. Molecular Therapy, 12(1), 171–178. https://doi.org...Researcher Perspectives on Data Sharing in Deep Brain Stimulation. Frontiers in Human Neuroscience, 14(December...

... science.                    Two of the platforms we use at Addgene: YouTube, for longer educational...octogenarians is going to be a hard sell!) Digital platforms often have demographic data of their users readily...geographic location. For example, not all social media platforms are available or used in all countries; not all...the limitations and potential of your preferred platform before you start can save a lot of trouble in ...requirements.  No visuals. Variety of hosting platforms can splinter audience. Primarily digital distribution...communicators. Well-developed content creation platforms.  Algorithms can make it difficult to reach people...

...GFP derivatives, but are instead related to the dsRed protein isolated from Discosoma sp. Similar work ...common fluorescent proteins and their mutations relative to wildtype GFP. Although not listed here, many... fluorophores Fluorescent Protein Mutations Relative to Wildtype GFP EGFP F64L; S65T EYFP S65G;...that separates mixtures of cells into distinct populations based on fluorescent signal. Thus, FACS can be...has also been used to identify particular cell populations in drug screens, to visualize micrometastases...

...receiving the translation, Addgene contracted with a professional scientific translator, L. Denise Roche...101 eBook is now available in Spanish!  This translation was gifted to Addgene by Daniel Mejías Sánchez...Roche, to edit the translation. We are pleased to announce that CRISPR 101: Una herramienta de edición ...  Many thanks to:  Daniel Mejías Sánchez, PhD, translator  Denise Roche, PhD, MD, editor, ldeniseroche@...

...deaminates a target cytosine (C) into a uracil (U) base. Later the resultant U-G pair is either repaired by cellular... (PSU) is adapting and improving base editing platforms for plant systems.  Adenine base editors for ...editing efficiency based on the location of the ‘A’ relative to the protospacer adjacent motif (PAM). ABE7.10...edit in a stretched window ranging from 1-17 bp relative to the PAM sequence (Zong et al., 2018) (see CBE... They can, but they require other base editor platforms. Hua et al. constructed a number of plant base...editing experiments in plants Choose a base editing platform depending on the location of your target base ...

...Transcribing and translating a protein fluorophore also takes time - bypassing translation altogether with...affinity, were eventually called aptamers, from the Latin aptus, meaning “to fit”. SELEX could be used to ... protein fluorophores and quenchers are still relatively bulky. One of the advantages of oligonucleotides... context of a paired fluorophore protein. Years later, Samie Jaffrey’s laboratory took advantage of their...hydroxybenzylidene imidazolinone (DFHBI) with relatively low fluorescence in vivo, the Jaffrey lab used...

...L-arabinose) without signs of toxicity or burden relative to a fluorescent protein control. We also added.../L yeast extract, and 5 g/L NaCl) or on LB-agar plates containing glucose and L-arabinose at 37oC for ... arabinose and colonies on arabinose-containing plates will be visibly white, and gas vesicles in cells...from left to right: colony whiteness on inducer plates, white inclusions in cells with GVs via phase contrast...heterogeneity required single cell screening to isolate a clonal cell line capable of producing ultrasound...homogeneity, as the cells could be batch-sorted to isolate a polyclonal cell line with strong gas vesicle ...

...the ratio of genome-containing viral particles relative to the total number of viral capsids (which can...vectors consist of full particles (white arrowhead) relative to empty particles (black arrowhead). Scale bar...measured by qPCR.   Physical titer is typically calculated by two popular PCR-based methods: Quantitative...is a factor in AAV titration since the starting template is the purified viral vector solution (which contains...be mitigated by using purified viral DNA as the template instead of the intact AAV particle. Finally, ...absolute titer values may not be as important as the relative titer values. Finally, because physical titer ...

...space — developing tools for cellular targeting. Lately, they’ve been working on two tools that expand ...safe to say the sample had their attention! They isolated DNA from the sample, amplified the CRISPR locus...most common RNA targeting system, Cas13, the "collateral" activity of the enzyme can lead to off targets...this information to identify a large variety of related bacteria with similar Cas systems. Some of these...knockdown efficiencies. The ability to identify related Cas systems opens up the potential of further development... also compatible with both AAV and adenovirus templates. The system, which is larger than Cas9, can also...

...cells with exogenous biotin. After a few hours biotinylated proteins can be captured on streptavidin affinity... on neighboring proteins resulting in their biotinylation. The advantage of APEX is its speed, labeling...biotin catalytic domain to allow for promiscuous biotinylation. Compared to BioID, BioID2 exhibits enhanced...ways to detect RNA-protein interactions that can regulate multiple biological functions. RapID (RNA-protein...Any proteins that bind the RNA motif will be biotinylated.  Find more details about RapID! EXCELL: Proximity... Plasmid 101 Protein Expression Plasmid 101 Biotinylation Other resources Proximity labeling review:...

...undergo post-translational modification with fatty acyl chains. For instance, myristoylation requires an...Also, in cases where the protein of interest is relatively small, adding GFP may alter diffusion rates. ...Although bioluminescent probes are usually dim relative to fluorescent probes, recent engineering efforts...an N-terminal consensus sequence (MG-) and prenylation requires a C-terminal consensus sequence (-CaaX... that the N- and C-terminus of GFP itself are relatively close, thereby minimizing the disruption of the...

...year. Feel the love with MTRIA sensors One of the latest additions to Addgene’s catalog is a new class of...domain of ZAP70 (ZtSH2) which binds only the phosphorylated state of an immunoreceptor tyrosinase-based...only when the RTK is activated and the ITAM phosphorylated. To demonstrate that pYtags can be generalized...smell the flowers - or rather, the lactate! The latest biosensor from Gary Yellen’s lab is LiLac, a fluorescence...concentrations of lactate. The microfluidic screening platform developed here is also a promising approach for...other targets. Figure 6: A) Schematic of BeadScan platform for screening candidate biosensor variants for...

...task would take longer than I thought, I’d be too late to meet up, and I’d just end up spending more time... friends, but didn’t realize it until it was too late. I didn’t spend enough time thinking about the possibilities...while the longer tasks will give you larger payoffs later. Don’t be afraid to accomplish some quick but moderately...assess whether that time was sufficient for the task later. If not, then you should definitely allot more time...friends and come back to the lab to split my cells later. Eating with friends has a somewhat immediate deadline...may have prevented me from splitting cells until late in the day anway. There are a few things I’ve found...