...about being a writer that can be a very painful process. And certain stories can make you feel like ‘I’m...shared with a postdoc, and I’m like, “Oh my god, what can I do?! This is a mistake! Why did I even come [to...that’s something that good storytellers in science can do.” Susan’s number one piece of advice for getting...communication: “Find as many opportunities as you can to get your name on things.” Susan personally found...series, but also being my first interview ever! You can follow her on Twitter @sejkeown. If you enjoyed this...Susan’s NEI internship. Application information can be found here Find tips for using social media as ... staff writer at the non-profit Fred Hutchinson Cancer Research Center. Susan Keown’s science communication...

... Always wear gloves, as the RNases on your hands can degrade RNA. Keep a clean work area, which may include...to remove residual DEPC. Alternatively, glassware can be baked at 180°C for at least 4 hours.   If your...layer. The top aqueous layer is extracted and RNA can then be isopropanol precipitated. This method reduced...samples cold until homogenization and cell lysis. This can be challenging depending on your laboratory situation...TRIzol® and ammonium sulfate stabilizing solutions can be made in house.   A common problem with no-kit ... extraction methods is the carryover of DNA that can potentially complicate results of a downstream application...expression. There are several things that researchers can do to combat this issue. First and foremost, be mindful...

...strong problem solving skills alone can take you - just because I can resolve issues effectively does not...problem solving skills” and look at how scientists can develop and highlight these valuable skills. Hopefully...found that taking a conscious or active approach can be hugely helpful not only for developing a skill...entire problem solving process on your own, but you can start by paying attention to how your mentors reach...solutions and ask them to explain their logic. Or you can try applying the scientific method to everyday problems...positions. Highlighting your problem solving skills can help convince a future employer that you are well...

...we learned how the Cre-loxP recombination system can be used to induce site-specific recombination events...flox your genes of interest, a powerful FLEx switch can be created for your specifc experimental needs. ...and lox511 are recognized by Cre but lox511 sites can only recombine with other lox511 sites, not with ...Once the SSR of choice is introduced, recombination can proceed either by first utilizing the loxP sites ... single gene deleted in your knockout mouse, you can design a FLEx switch conditional knock-in cassette...rescue experiment with a wildtype copy of a gene, you can also design FLEx switches to conditionally test the...orientation in the presence of Cre. A similar strategy can be employed for "Cre-off" (i.e. your gene of interest...

...need. You can find all the hot plasmids from 2017 below. With over 50,000 plasmids, we can't write posts... a valuable tool for the research community. You can find the AAV vectors for infecting GABA-ergic interneurons...control of enzymatic activity. In addition, proteins can be “caged” by fusion to specific domains thereby ...mCherry variants to the repository. These variants can be used as tools to learn about the influence of ...interest. NHEJ- or HDR-induced mutations in ATP1A1 can render cells resistant to the small molecule ouabain...detailed protocol at Protocol Exchange. Plasmid maps can be found at Addgene and in Figure S12 of the publication... ladders (aka DNA molecular weight markers) that can be used to measure the size of DNA fragments. Depending...

...touch, and be a more attractive candidate for that ideal job. How can you identify your transferable skills...bills and having a life outside my job. The same can be said about teamwork, communication, writing, management...owning your transferrable skills yourself so that you can grow them even more, get maximum satisfaction out...transferable skills: Put yourself in situations where you can learn more about possible strengths and weaknesses...that are just not being tapped at the moment? You can take this time to step up and develop your nascent...detail. If you are managing more workflows than you can count on your hands and toes, you’ve likely collected...in the lab, or organizing a research project, one can assume that you’ve had to flex your conflict management...

...parents who can answer some of the surprisingly technical questions elementary students can ask. It doesn... could contribute to the school, too. Though you can recruit parents from any background, it really helps... our math tables!  Be realistic about what you can teach  5-10 year olds in a single evening. Grades...STEM activity fun,” but rather “let’s see how we can make this fun activity relate to STEM.” Figure 1 ...teachers and other science communicators.  What can CRISPR do? I spent more time on this section of the...way to make this section more interactive. If you can think of a good analogy that involves student participation...engineering technology is moving along much faster than it can be officially regulated. I did not try to sugarcoat...

...and sgRNAs can be found here. Neuron-optimized viral vectors Alternatively, Cas9 activators can be delivered...However, using CRISPRa to target individual genes can be relatively laborious because multiple plasmids...expression. Constitutive or inducible Cre recombinases can be introduced by AAV or by mating with Cre-expressing...distinct transcriptional start site. Because CRISPRa can be targeted anywhere in the genome, it is relatively...BDNF protein. To demonstrate the specificity that can be achieved with CRISPRa, the Day lab singled out...The Day Lab’s neuron-optimized lentiviral vectors can be found here. High-throughput screening strategies...used by the Qi Lab for this particular application can be found here.  Why CRISPRa? Although cDNA overexpression...

...senior faculty members who can help you identify overly ambitious goals and can help you distinguish mechanistic...question, the methods, the tools or techniques that can help resolve the question, the anticipated outcomes...the validity of a single model. The first approach can lead to new directions, while the second approach...-like mini-organs or organoids, neither of these can substitute for a liver transplant. My lab will combine...problem. Creating a coherent unified story and model can help make your ideas easier to understand and more...outside your lab and outside your field. If they can understand and are engaged by your presentation, ... getting back to your story. Committee questions can derail your talk. You need to balance respectfully...

...with this rate varying across cell types. Variance can be caused by cell cycle rates; rapidly diving cells...available sister chromatid, though exogenous DNA can be used as well. Below are some considerations for...donors (ssODNs) with homology arms as short as 40 bps can efficiently knock-in a few hundred bps of sequence...guide recognition sequence in the donor DNA, you can guarantee there won’t be re-targeting after HDR. ...tilt the scales in your favor for an HDR edit, you can try one or more of the below suggestions. Disable... those cells! To take this one step further, you can inhibit cell cycle regulatory proteins responsible...are arrested too long. Prolonged cell cycle arrest can lead to apoptosis in as little as 10 hours, so read...

...License. Importantly, even commercial antibodies can be tethered on the viral surface, expanding the range... with fusogens from different viral families and can be used to target multiple delivery systems, such...lentivirus and MMLV gag, to specific cell types. It can also make CAR-T cell therapy more accessible by simplifying...cells directly within the body. Overall, DIRECTED can increase on-target delivery with various delivery...labeling and TransitID By Mike Lacy Proximity labeling can identify protein interactions or map a local proteome...different cellular compartments. They showed APEX2 can use an orthogonal substrate, alkyne-phenol, to label... slow-growing protein fiber in the cytoplasm and can be fluorescently labeled. Using lentiviral vectors...

...Plasmids need antibiotic resistance genes so that you can specifically isolate bacteria containing your constructs... division. These non-plasmid-containing bacteria can compete with and confound the goal of your experiment...transformation (the dreaded 60 minutes shaking at 37 degrees) can be shortened since the plated cells will have time...plasmid and its associated resistance gene. They can grow proximal to a colony that does contain the plasmid... benefit it offers is marginal. Fortunately, amp can easily be replaced with carb if satellite colonies...compared to streptomycin. Spectinomycin/streptomycin can be successfully used for plant and bacterial culture...genes in their genomes. If you need a plasmid that can go between plants and bacteria, it’s a great option...

...response to an experimental treatment can be monitored. This can reveal previously unknown functions for... specific tissues such as blood cells or neurons can be fluorescently labeled, allowing researchers to...mutagenesis screens. Microscopy-based lineage tracing can help illuminate how signaling pathways—such as those...abnormal cell types. Sequencing-based lineage tracing can help elucidate the gene networks that transform zygotes...neuroscience investigations. For example, neuronal activity can be tracked in whole brains of swimming, actively ...manipulation of brain activity: optogenetic approaches can be used to determine the consequences of activating...Lineage tracing studies highlight how these advantages can translate into insights in developmental biology,...

...with the addition of N-glycans [9]. Both of these posttranslational modifications can disrupt FP folding ...fluoresce. A dark population is not readily apparent and can confound quantitative imaging experiments or even...susceptible to these non-native modifications, which can all potentially affect function [5]. We and others...cellular function or possibly viability. FP misfolding can even disrupt fusion protein localization. For example...pathway, a dark pool that is mislocalized to the ER can be detected with immunofluorescence. This population...experiments, please consider the following: How can one ensure that an FP fusion with a non-cytosolic...inert green, cyan, yellow, and blue variants that can be used in various combinations. For example, with...

...to DNA extraction and quantification so that you can focus on what you want to do with your plasmid without...starting with 210 plasmids from the collection. You can find the full list here. Most of these plasmids are...cloning. Plus, we decided to include 10 plasmids that can be directly used in the research on the SARS-CoV-...out this service! Let us know what you think. You can also let us know which plasmids you’d like to see...

... you.” “Also a mentor can give you realistic expectations. (Don't think you can finish that book in 4 ..."The greatest good you can do for another is not just to share your riches but to reveal to him his own...experience. Navigating a fulfilling career in science can be challenging – is advice and guidance from only...Advisor/PI who will also be a good mentor? Where can I find a Mentor? What is this I hear about “group...

...light. PA-FPs can be activated from low fluorescent states to high fluorescence states, they can change from...fluorescent color to another, or their fluorescence can be switched on and off reversibly. This ability to...Irreversible photoactivation Irreversible PA-FPs can be switched on from a state of dim or no fluorescence...photoconvertible fluorescent proteins (PC-FPs). PC-FPs can convert from one fluorescent color to another (e.... is another monomeric PA-FP which unlike mEosFP, can form it’s chromophore at a more comfortable 37°C ...reversible PA-FPs (also known as photoswitchers) can be photoswitched from a dark state to a bright fluorescent...non-fluorescent state (“quenching”). The photoswitch can occur multiple times by activating with two distinct...

...base-to-base conversions (Anzalone et al., 2019). Plus, it can combine different types of edits with one another...extends the reach of CRISPR genome editing, as it can edit near to or far from PAM sites, making it less...availability than other methods. The PAM-to-edit distance can be over 30 base pairs for prime editing. Since PAM... (in certain circumstances) Current base editors can perform any base-to-base conversion in principle,...require multiple steps and enzymes. Prime editing can encode any of the 12 possible base-to-base changes...bystander editing is unacceptable, prime editors can be used to avoid this possibility. However, there... strategy, cell type, and goal of the experiment can all impose different constraints and affect how you...

...about how treatment regimens can be improved and how the therapies themselves can be made more effective. ...future immune surveillance. Dormancy of certain cancers can be attributed to the concept of immunoediting...equilibrium stage. This can be called “stalemate equilibrium” where the cancer stops growing, and the ...that eliminate early stage cancer cells, chronic inflammatory responses can augment tumor growth. Indeed...inflammation from infection can further divert the immune system from fighting cancer. Surgical removal causes...treatment strategies can be enhanced. Types of immunotherapeutics In lieu of the cancer-immune system interactions...because they can help start a sequence of immune reactions that aid in the fight against cancer. Sipuleucel-T...

...or A much further from the ‘NGG’ PAM cannot be editable? They can, but they require other base editor ...about CRISPR, one question probably comes to mind: can CRISPR be applied to fix the defective base precisely...CRISPR-mediated base editing (BE), these types of repairs can now be done more efficiently than before. A base ...guanine (G)? You may wonder if a CRISPR-base editor can perform this change. However, there are no naturally...successful evolution of an adenosine deaminase that can act on DNA for adenine base editing. They first created...pKABE6.3, pKABE7.9, and pKABE7.10) so that the user can optimize base editing efficiency based on the location...to the protospacer adjacent motif (PAM). ABE7.10 can edit target 'A' if it lies in a small genomic window...