...proteins are provided in trans (see Other Important Components of EBOV Minigenome System). However, minigenomes...sense RNA viruses (reviewed in 2, 3). Other important components of the Zaire ebola virus minigenome system...antiviral compounds which often target the viral polymerase. Technical tips It is important that all of...particular observation can be easily studied as each component can be examined independently. In addition, experiments...efficiently transfected into cells. All of these components are necessary to efficiently reconstitute a functional...utilize the minigenome as a template. Thus, it is important to choose cell lines that are highly transfectable...plasmid transfected into cells are also extremely important. For example, particular ratios of plasmids encoding...

...interacting proteins. No spatiotemporal control. Many PPI detection methods lack spatiotemporal control or an “...Split-BioID, a spatiotemporally controllable version of the BioID protocol. The critical component for both ... lysate fractionation. Important considerations for Split-BioID It’s important to remember that with Split-BioID...proteomics approach to monitor the composition of spatiotemporally defined protein complexes. Nature communications...this limitation by creating Split-BioID, a spatiotemporally controllable version of the proximity-dependent... other PPI methods, including BioID, is its spatiotemporal controlled labeling of interacting proteins...protein will be biotinylated. This makes it’s impossible to distinguish proteins that directly interact...

... methods. Across these methods, the following components are needed: γ-Retroviral transfer plasmid encoding...similar to lentiviral packaging methods. The three components described above (envelope, packaging, and transfer...γ-retroviruses differ from lentiviruses? There are a few important differences between engineered γ-retroviruses ...because once in the host cell cytoplasm, the viral components are small enough to enter the nucleus through...slightly different isoforms of these packaging components. Therefore, lentiviruses may not be efficiently...plasmid backbone is meant to be descriptive of the components found within the backbone. For example: pLXSN...particle containing Matrix, Capsid, and Nucleocapsid components. Pol Precursor protein containing Reverse...

...regulation, but enhancers and chromatin looping play important supporting roles. Enhancers are cis regulatory...of a stable cell line that co-expresses three components: dCas9 with an added biotin acceptor site BirA...genomic location of interest  When these three components are co-expressed, the gRNA targets the dCas9 ...with primary cells or rare cell populations. Key components of the CAPTURE 2.0 system To improve upon the... Using this tag reduces the number of CAPTURE components that need to be delivered to cells from three...lentiviral delivery of the two remaining CAPTURE 2.0 components eliminates the need to create stable cell lines...networks’ of 157 super enhancers.  3. Measuring temporal changes in chromatin interactions A third application...

... of 10 mL, we’ll want 200 mL of media total. Importantly, we’ll actually make a bit more than 200 mL (...

...Delivering CRISPR components to plant cells Once you have selected the correct CRISPR components for your application...deliver these components to your target cells. Remember, efficient delivery of CRISPR components is essential...organisms, some plant-specific changes to CRISPR components are necessary to enable genome editing in plant... academic researchers through Addgene. CRISPR components for plant genome engineering CRISPR can be used...genome of plant cells. The structure of the gRNA (composed of a ~20 nucleotide targeting sequence and ~75...line will result in a failed experiment. CRISPR components can be expressed stably or transiently depending...delivery method and cell type in question. CRISPR components can be delivered and expressed transiently using...

...genes important for mesoderm differentiation Now, let's turn tinman and bagpipe, two genes important for...protein Groucho, a transcriptional co-repressor important in development. Groucho family proteins are found...Dana Farber Cancer Institute! Sonic Hedgehog, an important regulator of embryonic development The largest...video game character but rather to an extremely important gene in animal development. To understand where...1997). We have a lot of plasmids that contain components of the SB transposon system including those from...but think again. Spaghetti squash is actually a component of the fly actomyosin cytoskeleton that regulates...regulates cellular contraction. This network is composed of actin and myosin which interact with each other...

...Starch-based compostable bags for kit components Recyclable Yes, all kit components and packaging...packaging Yes, most kit components and packaging Yes, most kit components and packaging Parts Sold...standby to fall back to? We have outlined some important considerations to help you choose the plasmid ...experiment), but issues with DNA quality can inhibit important reactions like PCR or restriction digests. Transfection...the micrograms scale and up). But perhaps more importantly, the DNA needs to be free of contaminants, such... very similar. However, keep in mind that kit components (i.e. buffers, columns, or beads) are designed...ordering department for these.  While budget is an important consideration, you do want to make sure you’re...

... do so have been hindered by a lack of tools, compounds, and assays to monitor activation of these receptors...to easily design, generate, and test constructs composed of complex genetic elements. To demonstrate the...protein-coding genes, and is unique in that it is composed of two sub-pools. The first is a base “90k library...and to deal with the increasing variety of new compounds being generated, there is a need for genetic optimization...identifying and studying interaction networks is important to detemine how these systems work. Researchers...identified through transcriptome sequencing. Importantly, results obtained using TRIBE compared favorably...auxotrophic markers. Read more about the Ralser lab's important new tool for the yeast community in their open...

...target genome. They did this by selecting the important elements from existing transposon and vector systems...Mitochondrial health, transport and turnover are important features to be monitored to study mitochondrial...of oxidation (dehydrogenation). MitoTimers are composed of a Timer protein fused to the mitochondrial ...cytochrome c oxidase subunit VIII, so that they can be imported to the mitochondrial matrix. Gottlieb’s lab has...mitochondria in the networks marks the differences in import of newly synthesized (green) protein and organelles...May 1;126(Pt 9):2102-13. Assemble multiple component plasmids from building blocks via Golden GATEway...existing elements. DNA sequences for a desired component are cloned into Golden Gate entry vectors via ...

...Stanford University. Sex and gender are critical components of biological research that are often forgotten...into research. Why are sex and gender analysis important? Doing research wrong costs lives and money. According... goal of gendered innovations It is crucially important to get the research right from the beginning. ...unique to male stem cells. Nor did they detect important differences in function between male and female...many factors that interact with sex, and it’s important to analyze these factors during an experiment....include sex in experimental design Recognizing the importance of analyzing sex, the National Institutes of ... tissues—when that information is reliable—is important to ensuring excellence in basic research.  In ...

...about it! Split Cas9 system The Cas9 protein is composed of an n-terminal DNA recognition domain and a ...self-assembly. In an effort to obtain even more precise temporal control of gene knockout or activation, the c-...)-FRB-NES). This system provides users greater temporal control over CRISPR/Cas9 mediated genome modification...activation allows scientists to spatially and temporally control which genes are turned on or off in a...principle of protein multimerization is seen in many important biological contexts, such as the enhancement of... Understanding intracellular pH regulation is important as pH regulation has roles in many cellular functions....0043072. Photoswitchable tools for spatial and temporal control of cell events Three years ago, Brian ...

...one receptor, so it is important to fully characterize the compound, or compounds, you choose to ensure...Targeted and specific delivery becomes especially important for studies in neuroscience where specific tissues...neuron need to be targeted. Targeting is also important in the development of therapies when expression...Picking Ligands Picking the right ligand is also important for building a chemogenetic system. Your experiment...cellular response you aim to manipulate. It is important to consider that the same receptor can elicit ...pharmacodynamic actions of clozapine-N-oxide, clozapine, and compound 21 in DREADD-based chemogenetics in mice. Scientific...

...weekend - why? Do they have an important plans? What is most important to each person? Someone has to...big conference - Is one of those options more important to you and why? What are their expectations/... you. Her feedback on your manuscript is more important to you than collecting your images this week. ...scenario where both people get something valuable. Importantly, as the third example shows, the ultimate resolution... the need to be thoughtful about what is most important to each person. It can especially be difficult... that partner with Addgene. What are the most important issues for our partners and how can Addgene work...differences is a crucial part of understanding what is important to people so that you can approach negotiations...

...Effects on protein function and cellular components: It's important to assess how dye-conjugation affects...enable highly specific visualization of cellular components, typically a protein of interest. Fluorescent... function and its propensity to bind cellular components. Some dyes target specific organelles like mitochondria... interpretation of experimental results. It's important to consider how dyes can influence receptor conformation...wavelengths for green, red, and far-red dyes is important for intracellular fluorescence live imaging ​​(...from cells under certain conditions, which is an important consideration for assay design (Breen et al., ..., 2016). When selecting dyes, it's important to test them under experimental conditions. Common issues...

...are the various tools and components necessary to run it? And, most importantly, what are the methodological...show causality in neural circuits with incredible temporal and spatial precision (I’m not going to discuss...Deciding how you want to use optogenetics The most important question to consider is what exactly do you want...cell type of interest is obviously of critical importance to any optogenetic experiment. There are numerous...of North Carolina) that offer his constructs. Importantly, control viruses (not expressing the opsin) are...for successful optical stimulation.  It is also important to note that some promoters are so robust they...next week’s post we’ll delve into some of the important materials science considerations for your optogenetics...

...of the independent variable you are testing. Importantly, they demonstrate the functionality of your experimental...non-independent variables.  To illustrate the importance of control plasmids, we will take a step beyond... the transfection reagents and process. It is important to note that optimal experimental conditions, ...Control (mentioned above) could also serve as an important negative control. In this context, the empty vector...interpretation of the results because it provides an important reference point when analyzing the specificity... same time. Although the replicate control is important, it speaks to the consistency of the reagents ...controls are more time consuming and ultimately more important. Ideally, each biological replicate should use...

...microtubule-cutting sword Microtubules are one of the most important components of the cytoskeleton. They provide cells ... if you’re interested in that other most-important component of the cytoskeleton – actin? Actin dynamics... example, microtubule-destabilizing drugs are impossible to target to specific subcellular regions, laser...manipulating Rho GTPases based on BcLOV4. These single-component systems take advantage of BcLOV4’s light-induced...powerful enough, what about altering the chemical composition of the membrane itself? The Baskin Lab previously...K. H., & Chow, B. Y. (2022). Designing Single-Component Optogenetic Membrane Recruitment Systems: The ...

...aligns with your interests. We've outlined some important factors to consider and summarize helpful advice...featured in "9 Tips to Achieve Success in Academia". Important factors to consider when choosing a research project... project After joining a lab, the next important decision to make is “What project will I work on?” In...typically require a diverse set of skills. It is important to know who in the lab and/or what facilities ...principal investigators Tom Ellis highlights the importance of keeping up-to-date with the literature of ...aligns with your dreams. He also highlights the importance of combining “new discoveries or technologies...will be fresh as well.” Don't underestimate the importance of choosing a solid research topic and approach...

...replicate the results of another’s work, it may be impossible to determine precisely how the methods were implemented...result after conducting a replication study. An important difference between the original setting and the...genetic variation, or different handling can all be important.  If we eliminate the problems above: unclear ...research finding we can be more certain that some important, previously unknown variable actually makes a ...all too often glossed over. This is why it is important to be able to look under the hood at the raw data... versus control, and you think there are four important times to measure that growth rate, you report ...everyone agrees that the research questions are important enough to be answered and the methods are appropriate...