...ICC) assays.  Figure 1: Three ingredients of epitope-tagging experiments. Cas9 protein, annealed cr:tracrRNA...procurement of cr:tracrRNA reagents, we scaled up these tagging experiments by assembling csRNPs in a ...set up csRNP knock-in experiments If your lab is new to CRISPR/Cas9 experiments, there is no need to worry...CRISPR reagents (turnaround time 5-7 days); because the tracrRNA is the same in all csRNP experiments, it...proteins. Similarly, in ChIP-seq assays, chromatin fragments that are bound by a protein of interest are pulled-down...adoption of CRISPR/Cas9 technology and rapid improvements in the field, it has recently become possible...CRISPR depends on successful delivery of three reagents to cells: a guide RNA, Cas9 protein, and a donor...

...receptors) or with experiments on permeabilized cells. Biarsenical labeling reagents If you want to label...control experiments are needed to establish the dynamic range of your assay. These experiments include...pressing issues in your laboratory, then FRET experiments are a viable biophysical path to answers. What...lipid bilayers, organelles or other cellular impediments. FRET has been used in many applications, including...fluorophore delivered to the right place? FRET experiments require targeting the donor and acceptor fluorophores...experimental design is that the targeting specificity requirements are different for the donor and acceptor fluorophores... on the same street as the donor), then FRET measurements are possible. In fact, even if the majority ...

...creation process. Each module of Fragmid represents a class of components, such as your CRISPR enzyme or Pol...module, are the components. Using the handy website, users can simply select the components they want for...module fragments on the Fragmid website.   Step two will ask you to fill in module fragments from ...able to select components that work with your design. Note that you can browse all components by clicking...Fragmid doesn’t have the components I want? You can make your own Fragmid components! Ensure your backbone...’ terminus, and selection markers. Need fewer components? A spacer can be inserted instead, allowing everything...everything to stay in the correct order. Most Fragmid components are available for request on the Addgene website...

...quantitative data for regulatory elements, which has not been available for plants. Addgene: Will the MoClo tools...Remember plants. Every time you look at your plate, think about how much that’s on it are plants, and the...produce constructs and edit genomes so as to make plants, and agricultural crops in particular, resistant...elucidate the molecular interactions that go on between plants and their pathogens. As Patron explains it, her...been focused on gene transfer, from transgenes to plants, chloroplast to the nucleus, or pathogens to their...have tested them to show that they work well in plants and give similar stability to transcripts as the...construct, you don’t have to worry about repeating elements anymore. The kit contains sequences for several...

...science teachers reached out to their students’ STEM-oriented parents looking for experts to come give guest...can recruit parents from any background, it really helps if activities are run by parents who can answer...groups of about 50 students each, and each group went on its own wonderful tangents. Some groups asked... the annual Math & Science Night, a chance for parents and other members of the community to share their...general meeting and a means to request funding for events, consider reaching out to it. PTAs have more flexibility...event-planning base and will know how to reserve space, time events, and network for volunteers. Speaking of volunteers...you live in a school district that has a lot of parents in STEM careers, chances are they’re wondering ...

...scientific community represents the culmination of many such experiences. For example, students in Provost’s ...sciences by making a greater number of students active participants in the research process – so if this...provide the same kinds of opportunities for his students. But he also found that his teaching responsibilities...decided to adopt a more focused format, where all students work on malate dehydrogenase (MDH), a citric acid...ultimately settled on a two-semester model, in which students learn to express, purify, and analyze MDH in semester...skills to design and carry out their own novel experiments in semester two (Knutson et al., 2010a,b).    ...research activities in a CURE must be authentic – students should be attempting to generate new knowledge...

...recombinases, regulatory elements, developmental regulators, and lots of viral components. This kit is also ...bacteria, protists, plants, and animals (Chiasson et al., 2019). Many plasmid elements such as promoters...systems, mammalian cells, yeast, flies, and even plants! In this blog we will review common uses of MoClo...chemical synthesis of these molecules. To make large amounts of pure product in an affordable way, researchers...Pérez-González et al., 2017). In addition to additional elements for S. cerevisiae chromosomal integration and ...great starter for your general plant cloning experiments. Addgene also has two additional MoClo plant ... a variety of Cas enzymes with different PAM requirements and editing abilities, such as base editing,...

...are too few resources to teach students how to perform these experiments in the beginning. In most classrooms...lab publication [1] so that the students performing these experiments have a good baseline for comparison...techniques such as basic observation and dissections. Students don’t usually perform more modern techniques used...in genetics or molecular biology because the experiments are either not accessible or too challenging ...easier to perform advanced developmental biology experiments in planarians with their recent plasmid deposit...deposit. These plasmids enable two main types of experiments: Whole mount in Situ Hybridization (WISH) for... expression. The 49 deposited plasmids enable students to study gene expression and functions of key ...

... sequence in between two fragments of EGFP in pCAG-EGxxFP. The EGFP fragments contain 482bp of overlapping...between two non-fluorescent split GFP fragments. When the two fragments, expressed in the presynaptic region...genome. They did this by selecting the important elements from existing transposon and vector systems and...an all-synthetic vector that included only the elements needed for function. The lab validated this vector..., particular for those involving recombination elements such as FRT or Lox sites, and provides a modular...system for easier exchange and re-use of existing elements. DNA sequences for a desired component are cloned... Gate cloning step to assemble the individual components into a destination vector in a predefined order...

... reliable reagents for future research. I’ve talked to many researchers who have complaints about the ...provide you with additional validated reagents, and, with these reagents in hand you’ve got a good foundation...tries 5 separate experiments at once to get confirmatory data for a hypothesis and presents only the results... Scientific reproducibility As Jeffrey Flier points out in his talk (1:24 - 29:31), there are a variety...Freedman et al 2015). Finally, as Pamela Hines of AAAS points out on the panel, irreproducibility undermines ... 1: Inability to repeat previously performed experiments Researchers should be able to follow one another...another’s protocols and use one another’s reagents with the confidence that they aren’t simply doing an ...

...60.2%) 20 graduate students (25.6%) 12 masters students (15.4%) 46 (59%) of these students were in the last...-program self-assessment, participants completed follow-up self assessments and program evaluations every...self-assesment. Achievements directly attributable to participation in the program included improvements in writing... got a physical award, all the students experienced great improvements, not necessarily quantifiable, ...mentorship during undergraduate and graduate studies. Students also need more access to reputable, organized ... networks connecting international experts and students. In response to such needs, the International ... BWH and providing career mentoring to several students from Spain and all over the world, she thought...

...particular, two recent events have the potential to influence both graduate students and postdocs broadly... Board to allow graduate students who work as teaching or research assistants to join unions. The topic...example, Yale graduate students filed for union elections in only 10 departments across the university....flexibility to address the unique needs of students from different departments across the university and alterations...advocacy skills “bootcamp” for a focused group of participants. More information can be found at http://futureofresearch.org...F1000Research, 2014; Dolan, F100Research, 2016). These events have also started progress towards solutions. The...which several projects were initiated. Recent developments promise to further change the landscape facing...

...works in the lab knows how expensive tiny amounts of reagents are, how complicated (and again, expensive...FD-CF pellets are easy for students to use in the classroom because the students simply need to add water...Most importantly, experiments for FD-CF reactions can be fun and engaging for students. In our BioBits ...example, we’ve designed FD-CF experiments that result in outputs that students can easily sense, whether ...consider using some elements of BioBits to transition your undergraduate students from the classroom to...had this kind of experience earlier! After all, students build circuits in physics class to learn about...funding and the resources to provide them to their students. My desire to create a set of low-cost, easy-to-use...

...minimal upfront requirements and allows you to expediently join multiple synthetic fragments, create multiple...refers to a method of physically joining multiple fragments of DNA to create a synthetically designed DNA ...multiple mutations in one or multiple fragments, and generate constructs for producing single-guide RNAs... yielding scarless DNA assembly, and allowing fragments to be taken directly from restriction enzyme digests...the absolute end of a fragment. This means that fragments (containing regions of homology) cut from plasmids...planning. This cloning methodology requires that the fragments to be joined contain regions of homology at, or...homology can be contained within the individual fragments themselves, but doesn’t necessarily have to be...

...well treatments are working to fight the cancer.  Beyond early modifications to standard treatments and...are running out of hope with more traditional treatments, such as surgery, radiation, and chemotherapy...sufferers can see great promise in up and coming treatments like immunotherapy. Mesothelioma Mesothelioma...and late discovery mean that most mesothelioma patients have an extremely poor prognosis. Together, the...disease with only about 3,000 new mesothelioma patients diagnosed every year in the US. Therefore, mesothelioma...many forms of cancer, the primary conventional treatments for mesothelioma include surgery to remove cancerous...cases the cancer is diagnosed so late that these treatments do not work very well. The best option in such...

...failure to clearly articulate one's talents and ideas. Some key points from my ebook, "Applying for a Faculty...success is publications and grants. 2. Write succinct and easy to understand documents A common misconception...example documents. This part has generated a lot of positive feedback. Many of these documents are too...sections, along with new thoughts on research statements. Snapp, who is always looking for ways to improve...improve his eBook, adds: "If you have any questions, comments or suggestions, please do not hesitate to email... members will NOT be in your area. Therefore, documents need to minimize jargon. Equally importantly, ...seminar See item 2. Attendees will include grad students (that are solicited for feedback), possibly the...

...department worked with students to maintain these time goals during COVID-19? How are students funded? Are there... outside of just the program requirements: by talking to current students about the work environment and...and standards for hybrid, virtual, and in-person events change seemingly daily (and are inconsistent between... you’re applying to a school that requires its students to do rotations in different labs before selecting... universities offer free practice sessions for students doing virtual interviews, so check if yours does...keep yourself organized. By writing down your key points and keeping them at-hand, you can quickly reference...universities typically invite a select group of students to continue the application process through a ...

...compatible with all your components. Next, consider the best form for those CRISPR components based on your model...introducing those CRISPR components into your target cells. Delivery of these components into mammalian cell...considered during the planning process. For CRISPR experiments, the following framework can help get you started...most effectively achieved with different CRISPR components. Select the appropriate CRISPR tools for your...will you be editing? Do you want to deliver the components as DNA, or would mRNA or protein delivery be ...of the first steps is to identify what CRISPR components you will need to deliver. Minimally, one or more...model system. CRISPR reagents can be delivered via transfection, nucleofection, viral transduction or ...

...green (v857) variants differing in kinetics, affinity, and ∆F/F0. Introduce the two variants in three different...The new iGluSnFR3 variants should enable scientists to robustly detect signaling events in the brain and...models. Furthermore, they created three affinity variants – A184S (increased affinity for glutamate), S72A...iGluSnFR had limitations: they were not optimal for experiments requiring high signal to noise ratios, imaging...imaging of high frequency glutamate signaling events, and accurately differentiating between synaptic vs. ...Kaspar Podgorski’s lab have recently made iGluSnFR3 variants that are more sensitive and better localized for...predecessors. The detailed characterization of the new variants is available here. Upgrades to iGluSnFR3 To improve...

...split into two fragments that individually lack fluorescence. When complementary FP fragments are fused to...construction Fusing FP fragments to your PIPs requires careful selection of FP fragments and, preferably, some...that underlie protein function. However, FRET experiments can be difficult to implement in practice because...hypothesized protein interaction partners (PIPs), the FP fragments form the complete FP because the PIPs they’re ...small-molecule inhibitors of a particular PPI. In such experiments, small-molecule inhibitors are tested in parallel...Design a BiFC Experiment Much like FRET, BiFC experiments must be carefully optimized to be successful....will allow you to determine where to fuse the FP fragments. In the absence of this information, you can fuse...