...less phosphatase-accessible. Kinase Translocation Reporters (KTRs) Kinase Translocation Reporters (KTRs...fluorescent kinase reporters. A KTR is comprised of a fluorescently-tagged substrate for a kinase of interest...proxy for kinase activity.  Advantages of KTRs When compared to FRET reporters for measuring kinase activity...Kinases: they regulate many proteins, with ~1/3 of human proteins predicted to be phosphorylated on at...for regulating signal transduction and measuring kinase activity at the single-cell level can aid in drawing...phenotype. One method for monitoring live single-cell kinase activity is FRET, but FRET reporters are challenging...Covert Lab provides an alternative tool with their Kinase Translocation Reporters (KTRs) whose cellular localization...

...am currently working on projects for the Science Communication function at Addgene. Working a 9-5 As it...one semester (the majority spend two to three) working in your chosen field. This cooperative education...bioengineering student, to Addgene. I am currently working on the QC/QA team as a DNA Sequencing Intern. ...interview was intimidating to me, as I had a hard time thinking about how I could fill this time. I was relieved...environment on my team and at Addgene, I felt comfortable asking questions without fearing they would be disregarded...addition to these core hours, we have flexible working hours. For Addgene that means that it is possible...

...story. This year, we made significant strides in making science more accessible and reliable. We continue...flexibility. But this is just the beginning. We’re working toward giving you more choice in how you receive... every breakthrough enabled brings us closer to making an impact on humanity's greatest challenges. Thank...

... such as nutritional status. Within the animal kingdom, this information is relayed through factors (both...such as genes involved in metabolism, growth or cytokine action). Primary or core coregulators (those that...cellular roles. In the current phase of NURSA we are seeking to compliment these consortium-generated datasets...with a one-stop shop for plasmids and vectors, and making them available to all researchers for a reasonable...

...side conversations within the team. We are not talking about gossip here – this means encouraging a culture...Connie Cepko to learn about her management style.   Seeking feedback frequently One of the most important paths... only way.   From a colleague: MBWA "Manage By Walking Around" – don't hide in your office. Stop by instead...

...as-is, saving time and effort but risking the potential of them not working, or spend a few weeks designing...If you’ve ever been looking for just the right CRISPR vectors on Addgene and found instead ones that ... the heavy lifting out of CRISPR vector design, making it easier to get the vectors that are just right...right for your experiment. As always, we recommend checking your cloning via a miniprep and restriction digest...design. Note that you can browse all components by clicking the blue text in the upper right-hand corner.  ...Fragmid website.   If the vector is to your liking, your last step, Step 4, is to give the assembled...six or seven components — in most cases, that's making two or more plasmids — it may be more efficient...

...stories, and other 20th anniversary celebrations, by checking out our blog, social media, newsletter, and website...typically adopted by for-profit companies, while making use of our nonprofit status to achieve mission-...mission and work. In 2024, we will be actively seeking ways to support a global community where every ...the Addgene blog 2022 and 2023: Reflections and Looking Forward...

...Protein kinases deliver messages to the next members of their pathways, but where the kinases are located...subset of kinases. The Problem:  There is no common method for creating optically controlled kinases. The ...design for creating photoswitchable kinases and use MEK1 as a model kinase. In this system, light is used ...photoswitchable kinases: psMEK2, psRaf1, and psCDK5. MEK2 is a serine/threonine kinase with a similar ...photoswitchable kinases good for? There are a number of potential applications for photoswitchable kinases.  Below...Zhou et al. 1. Testing kinase inhibitors Zhou et al examined the ability of kinase inhibitors to alter Raf1... by intermediate messengers. Optical control of kinase activity can provide greater spatiotemporal resolution...

...jobs where all I did was public speaking. If you're also nervous, try taking a few deep breaths. Then, identify...breathe!)   Public speaking as a transferable skill The worst thing about doing public speaking as a scientist...Public speaking is not just one of the most common phobias–it’s a key part of being a scientist and an...support for your ideas, invite others into decision making processes, and even interview better! As a scientist...science communicator, I’ve done a lot of public speaking, and I'm happy to share what I've learned along...When my mind went blank, my options were to stop talking until I could present something coherent or to ...me accomplish my goal.   As a scientist, public speaking isn't a performance art. You’re (usually) presenting...

...accomplishments from 2022 and share what we’re looking forward to in 2023 and beyond. Last year, we distributed...rapidly changing environment. In particular, we are working to understand what challenges remain in materials...changing landscape of science. We are therefore embarking on a strategic refresh to understand this new ...help a great number of people. I look forward to working with Addgene colleagues, collaborators, partners...

...relationships because networking has gotten a bad reputation (as in…”I just hate networking”). Scientists enjoy... having to talk with strangers. Be bold in asking all kinds of people to connect with you and to share...the nonacademic sphere where most of us will be working.  Check out Joanne's Reddit AMA It should be no...fine to ask about specific jobs (that is what networking is about), but be upfront about interest in a... yes and they might say no but do not confuse networking for a job connection with ongoing relationship...relationship building through Informational Interviews. Making Your informational interviews productive Informational...remote conversations. In this interview you will be asking the questions, so be prepared. There are many good...

...definitely recommend checking out GCE4All’s website and resources, and then checking out all of the plasmids...focus on development, along with the inherent trickiness of using and optimizing GCE, led to a large toolbox...reagents!) for the use of GCE. In short, they’re working to make GCE available for all biomedical researchers...

...Taking the road less traveled and generating a knock-in cell line instead of a knock-out? We’ve got you... events will generate these types of mutations, making it an ‘easy’ editing job to get done. However, ...observed when inserts are within 10 bps of the break, making the cut off essentially as part of the templated...

...as coming from networking at the time, but that means it was the best kind of networking! Since nearly ... parents looking for experts to come give guest presentations. If you’re a scientist thinking about reaching...was very much in the making. One of the exciting, and challenging, aspects of talking to 8th graders was...my community. This past year, I spent some time thinking about how I could combine all of these roles by...space, time events, and network for volunteers. Speaking of volunteers, you can’t run an event like this...call to local parents who would feel comfortable speaking on topics that would compliment the STEM curriculum...been aimed at grad students, postdocs, and other working scientists, creating a presentation for 8th graders...

...gene expression of cells at a single time point, making it difficult to investigate how early expression...and cell identity at the single-cell level, thus linking ancestral gene expression patterns with reprogramming...reprogramming outcomes (Biddy et al., 2018). Tracking lineage and cell identity at the single-cell level with...re-expressed fibroblast genes. Most cells ended up taking the ‘dead-end’ path. The CellTags revealed that...intestine, demonstrating its utility in long-term tracking in vivo (Guo et al., 2019). Key takeaways of CellTag...cell-indexing tool that combines scRNA-seq and lineage tracking to track the dynamics of gene expression and clonal...cells or the clonal evolution of cancer. If you’re looking to use CellTag, you can find the CellTag libraries...

...Streaking for single colonies is an integral part of any bacteriologist’s skill set. So when Dave Westenberg... 1. Lab B needs a redo :-( @SandTBioSci #CompetitiveStreaking @Addgene pic.twitter.com/mk44J4Gcj9 — Dave... suspension,” says Westenberg. In addition to streaking for single colonies, these chromoprotein-expressing...

...when you can, instead of just asking for favors.  Don’t focus on networking with “important” or senior people...helpful who you’d like to get to know better. Networking is not about how many contacts you have; it’s...become a volunteer). Naturally running into or working with the same people is often the easiest way to...get a response. It breaks an important law of networking--don’t be creepy. There is a person on LinkedIn...they don’t know me. It’s a waste of our time.  Networking connections can last years This is a graphic ... the Addgene blog Browse other blog posts on networking Attending a conference soon? Get tips here Learn...

...founded in 1944 with the goal of facilitating networking between chemistry students, especially those ...scientific disciplines, as well as social and networking events. Imperial Synthetic Biology Society, ... synthetic biology by providing education and networking opportunities for society members as well as ...in society and to support iGEM alumni-industry networking to promote the creation of start ups and biotech...students and the general public, career events, and networking meetups. The society hopes to help young scientists...touch! Resources at Addgene Read Our Posts on Networking Get Career Advice from Addgene Bloggers Learn... networking...

...of cells. Looking ahead What’s next, you might wonder? Verkhusha says they are always looking to improve... If you start poking around on Addgene’s Fluorescent Protein Guide to In Vivo Imaging, you’ll pretty ... and intensity allows for optical labeling and tracking of proteins, organelles, and living cells in a...improve upon existing genetically-encoded probes by making them brighter, more photostable, smaller, and less... the 750 to 800 nm range, and his team is also looking in the direction of biosensors capable of revealing... muscles work inside a living animal. For those making use of these probes, he says, when in doubt, read...

...about replying and thinking about good questions and answers. Professional networking groups will often...Check out Joanne's Reddit AMA Who should I be looking for? Be on the lookout for friends or colleagues...who gives a great talk, ask them for advice on speaking. If you see someone with a great professional ...t doing it at all yet) watch my webinar, “Not Networking 101-Building Relationships for Success”. Choose...Choose 1-2 tactics to try. One of my favorites is making the pledge to have lunch or coffee with someone...started with mentoring. Read this Catalyst Report “Making Mentoring Work” to learn about what makes a good...to organize a successful peer mentoring circle. Asking someone to be your mentor Mentoring is a “loaded...

...editing and later looking for songs and sounds to synchronize with the images thereby making the videos more...contributed by guest blogger, Dalila Cunha de Oliveira. Bricking Science is an idea built, literally, 'brick-by-brick...Running Smoothly and "Gnome-Proof" This is why the “Bricking Science” YouTube channel was born. The name came...with each other through globalized, collaborative thinking. LEGO and science: The potential is endless One...feet), to connect people with science. And so, “Bricking Science” was born. We chose the stop motion format...approachable for LEGOs. Stop motion consists of taking many pictures, one for each position in a movement...video takes an entire week of our free time. After taking almost a thousand pictures, my husband Enzo will...

...2 Here! We'd like to thank Niroshi and Ben for taking the time to sit down and talk to us and wish them...

...genome randomly, while others use attP sequence docking sites so that you can control exactly in which ...at different loci. Fortunately, there are many docking sites available for integration. 2. How soon would...would know beforehand on which chromosome your docking site is located). This amounts to a small difference...important to note so that you know what to look for. Making your transgenic fly and downstream experiments ...experimentation. Drosophila are amenable to many different kinds of experimental assays depending on your interests...University. He has had nearly a decade of experience working with Drosophila as an experimental system. Jon ...Central PMCID: PMC1526508. 2. H. Jiang,  et al. Cytokine/Jak/Stat signaling mediates regeneration and homeostasis...

...importantly, this knowledge will stop you from embarking on a fruitless journey of trying to get males ...that. Now you have the basic knowledge to start working with flies. Like Addgene, the Drosophila community...chromosomes to use. If you’re not ready to start working with live animals yet, Harvard’s DRSC/TRiP- Functional...University. He has had nearly a decade of experience working with Drosophila as an experimental system. Jon ...

...has been supported by a Steering Committee and working groups with representatives from all areas of the...

...group in industry? The collaborative nature of working in industry and how delegating can empower people...she sets metrics for success. The importance of thinking about the big picture and not just the next experiment...Chapter 6: Advice to scientists who are planning on working in industry. The importance of recognizing and ...

...here and you can find more tips and protocols for working with lentivirus on our blog and protocol pages.... love to hear from you if you have used or are thinking about using our viral services. Please send any...

...mouse and human kinomes. While any subset of the genome is limited by definition, the kinome is a well-annotated...genetic screening project can be a tremendous undertaking, producing a wall of results that can only be... About CRISPR Pooled Library Screening Why the kinome? While properly designed perturbations – in the...incremental approach is prudent. What's in the CRISPR kinome library? Thus, we are glad to provide CRISPR libraries...pools provided in two sets, each with 4 gRNAs per kinase, so screens can be performed with either 4 or 8...individual lentiGuide gRNA plasmids targeting the human kinome. These plasmids contain the same 3-4 gRNAs found... Additionally, this set can be used for cherry-picking individual gRNAs that score as hits in pooled screens...

...need to be cost and time efficient. We do this by making our own competent cells and using a little-known...efficiency of transformation (for example, if you are working with a library or an especially low concentration...will work just fine.  A number of protocols for making competent cells are available online. A few examples...

... I found Vectr to be fairly intuitive and I was making figures quickly. Inkscape Inkscape is the most...images. For example, by selecting “Tools” and then clicking the “Usage rights” dropdown menu, you can search...

...fluorescent proteins to oligomerize is known as “stickiness.” Since the fluorescent tag should operate as... per unit.  There are several ways to measure stickiness. In vitro, ultracentrifugation or gel filtration...oligomerization in cells. In addition to measuring stickiness, several researchers have also collected data...characteristics are important, but have less priority than stickiness. In my experience, fluorescent proteins that ...independent of GDI-mediated extraction and actin-based trafficking." PLoS biology 13.4 (2015): e1002097. PubMed ... measurement of Gαi1, Gαi2 and Gαi3 activation kinetics in single cells." PloS one 11.1 (2016): e0146789...

...differently if I would be speaking to a colleague in the office next door. So be kind, helpful, and thorough...

...really fit in anywhere and everyone who was doing the kind of work my lab wanted to do already seemed to know...graciously took me for coffee and spent an hour with me talking about the tools I could use, and some experimental...This was invaluable to me as a bench scientist working at an institution where training opportunities ...network at the meeting. We found our mentors by asking for help with the program on social media. Many...between mentors and Singletons can accelerate the networking timetable for the Singletons in their bins.  ...organizers in their town hall.  Iterative improvement - Making Binning Singletons better Our experience running...leaps. Binning Singletons is its own organization, working to make scientific meetings more productive for...

...University. In this second post in our quick guide to working with Drosophila, you’ll learn how to maniupate ... YFG. You could compare this mutant fly to one lacking the mutation in the same genetic background. In...express YFG in a new tissue type (more details on making a fly line in the next blog post). For example,...get the right cell type expression as opposed to making a new fly with hedgehog under a new promoter each...University. He has had nearly a decade of experience working with Drosophila as an experimental system. Jon ...

...where I spent a couple of days talking to researchers about their work, picking up plasmid deposits, and trying...spend a large amount of my time meeting with and talking to graduate students, post-docs, and PIs. I won...enterprise work better and because I really like talking to people about and communicating science. The ...research at BU, I realized that I’m energized by talking to people about science and that I love explaining...running away screaming, but instead can involve walking alongside it and giving it a high five. While screaming...technical writing, teaching, presenting, analytical thinking, time management, information distillation, etc...transferable to many different fields. If you start thinking about preparing yourself for a career while still...

...business of iPSC clone picking, a 100-fold decrease is the difference between picking 100 versus 10,000 clones...efficiency.   But what’s better: PAM-blocking or guide RNA blocking mutations? This depends entirely on...efficiency of guide RNA blocking mutations. So if the locus does not allow blocking the PAM sequence (e.g...intended mutation while taking advantage of the improved efficiency provided by blocking mutations. Essentially... incorporation of the blocking mutation. Despite the fact that both the blocking mutation and our intended... same CRISPR/Cas-blocking mutation but not the intended mutation. Here, the blocking-only templates will...used CRISPR/Cas9 extensively for the purpose of making sequence-specific changes in human induced pluripotent...

...experience working in a developing tropical country. This is why I set up a placement working in a rural...work-life balance compared to working in research. My previous experience working in Industry was really valuable...sustainable. After working at this company for several years, I was ready for a change.  Working abroad: Research...new science graduates and the pro’s and con’s of working in different sectors. Luckily, I have worked for... decide for themselves.  Here is my experience looking for the perfect job. Summer work experience in... explain why I changed jobs. I highly recommend taking on a similar position in your field of interest...and problem solving. I believe that is something lacking during degree courses; they never really prepare...

...insufficient blocking) or reduced signal (too much blocking.) If you’re having issues with your blocking, first...it, try making fresh running buffer. Old or improperly made running buffers can cause all kinds of odd ..., degraded or improperly mixed blocking buffer, or insufficient rocking during incubation steps.  Many...pattern in your background, try asking more experienced labmates or looking it up online. You’ll find many...ResearchGate’s western blot forum.  Blocking Don’t skimp on the blocking time! Fill the time with smaller...  Troubleshooting Blocking reduces the amount of background in your blot, so blocking issues can either...you can try either longer blocking times or higher concentrations of blocking buffer. If you see a low ...

...room, so it is a good idea to start thinking about public speaking early and often. Two of my jobs have...supposed to be done. They will start looking at their phones. Speaking longer just makes you seem unprepared...choosing controls, or mixing clear solutions—public speaking is skill that any scientist can learn.  Any time...how to get it, especially if introverted ), and networking (how to get people to remember you). What better...Watch other people give great talks We are not talking about lab meeting here. Lifeless presentation of... Series. Watch how these speakers talk without looking at their slides or notes. Listen to their stories...and policy content. Watch Joanne give her "Not" Networking 101 Talk 2. Tailor every talk to the occasion...

...46 : How Kingdom ended up working in the life sciences and agriculture 5:47-9:35 : Kingdom's biotechnology...: Intro 1:01-3:48 : Kingdom Kwapata's background and education 3:49-4:35 : Kingdom on opportunities to...14:53 : Kingdom's current projects producing transgenic crops 14:54-23:44 : Follow up Kingdom's PhD projects...Seeding Labs is helping advance Kingdom's research 28:08-END : Kingdom's Thoughts on his time in Boston...family in Malawi that pushed science education, Kingdom Kwapata grew up to be one of the best of the best...Biotechnology at Michigan State University with the backing of a prestigious Fulbright Fellowship. Dr. Kwapata...research at Michigan State University 9:36-10:29 : Kingdom's transition back to Malawi and Lilongwe University...

...presenting on building relationships (also known as “networking”), one of the most common questions I receive...starting to see the advantages of using social networking to start, build, and track professional relationships...professional contact information available. When I am speaking at a new institution, for example, I will often...d be much more likely to say  “Hey, thanks for looking me up on LinkedIn!” resulting in just a little ...inviting someone to connect, perhaps you should be asking yourself if it’s really a good idea.  Pro tip: ... unless you’re on a profile page. You might be looking at a list of suggested connections and you’ll see...complete strangers as it borders on my first rule of networking--”If it feels creepy, don’t do it.” It’s just...

...chat box and upvote If the thought of walking up to the mic and asking your question in a large conference...in case someone’s internet cut out. “ On networking Networking can be more difficult in the virtual format...organic networking,” she says.Depending on the meeting, there may be a “place” for networking within the...answered this way and the hurdle of getting up and asking in front of many people is lower,” says Bachle....,” says Zhang. “Usually the moderator starts by asking the speaker the highly upvoted question.”If the...in the chat box. That’s another benefit to note-taking: you can revisit the Q&A in the chat box. Presenting...the participants to learn about what they were looking to learn in the workshop so they could steer their...

...scientist who began working remotely recently? Most of us at Addgene began working from home last week... mindfulness group is sticking with their Headspace practice even while working remotely. Social distancing...week to flatten the curve, but working remotely can be more difficult if your work is based in the lab. ...sharing links to online yoga classes. Get outdoors. Taking a walk or a bike ride is a great activity for both...the desire and ability to, some communities are looking for bench scientists to assist in labs that are...volunteering to foster a pet, for example. If you are looking for some resources specific to self-care during...check out Care For Your Coronavirus Anxiety and Taking Care of Your Behavioral Health. Work on personal...

... to DNA, crosslinking may not be necessary. This is known as Native ChIP.  If you’re working with a tissue...analysis process. And ChIP itself begins at...  Crosslinking Once you have your sample, the first step is...sample processing and therefore will require crosslinking, which uses formaldehyde or UV light to create...complexes together until you want to break them. Crosslinking is necessary for most ChIP experiences, but ...determine concentration.  Protein Capture After crosslinking and sonication or enzymatic digestion, you’re... not be a good choice) and the amount of cross-linking can affect epitope availability.  Collect the DNA...then incubate with Proteinase K to reverse the crosslinking. Finally, purify your DNA with a PCR purification...

...time for me to start considering my future path. Talking to Addgenies has helped me get a much better idea... Addgene’s Director of Product Management Eric Perkins, who has been in many roles in Addgene since he... really get me where I haven’t gotten myself.” Perkins noted that there is a more emphasis today about...those doors to be opened.” Addgenie Eric Perkins presented a poster a few years ago at a CRISPR ...away. Shannon Rinaldi, Addgene’s Lab Manager, was taking classes at her local community college when she...assistant in a lab at the university that I went to. Making that switch from being a grad student to an employee...Technician, dreamed of becoming a doctor. After taking a year break in between undergraduate school and...

...there's a big office vs. lab culture shock). I'm looking to see if there are any resources that can help...and many of my scientist colleagues have as well. Making the transition from the bench can be jarring for...aren’t a lot of resources out there for scientists making this transition. In this installment of the Addgene.... What causes the lab → office culture shock? Working in an academic research lab involves a few key ...your family. I talked to a graduate student at a networking event recently who referred to her lab as her...applicant asked if she would partake in “critical thinking” in the position to which she was applying. I ... stretch of time that doesn’t involve critical thinking. Former Addgenie Jessica Welch says, “[It] seems...

...researchers do learn MANY transferable skills while working in the lab and by offering advice on areas where... cell culture, I would often respond to friends asking me to dinner saying something like, “I’ll be there...ve learned a lot since my days as an undergrad working with cancer cell lines. As I started planning out... this post useful. After a bit of research and thinking about my own processes, I’ve broken down time ...until it was too late. I didn’t spend enough time thinking about the possibilities and how to prioritize ...the day, month, or even the year, you should be thinking about two main things: How useful the task is...time it will take you to complete the task When picking which tasks to actually do, you should err on the...

...afford them). I am talking about making sure the audience is interested, educated and talking as a result of...moderator should have a well-planned approach to making the event interesting and engaging. Long before...the pre-event interviews, the moderator should be taking good notes and discovering interesting stories,... panelists have the correct date, location and parking information and remind them about the event as ... a really lively discussion, I strongly suggest asking the panelists not to use any slides for the event...prevent any one panelist or audience member from taking over. My own personal rule is that I do not let...chose good panelists, they are going to be good at talking, so they will easily take 5 minutes to introduce...

...conference. Networking All of the things we've mentioned above could be considered networking, but here ...tips. Before the conference Networking is key even if you are not looking for a job - those connections...the elevator pitch, talks, the poster session, networking, and social media. The elevator pitch Before...poster or talk at the conference, you can end by asking them to come to your poster or talk. After the ...bored, or particularly excited about? If you tried tweaking your pitch throughout the conference, what worked...the poster session, send them a follow up email thanking them for their conversation and connect with them...social media. Social media Before the conference Networking can be made easier through social media. Use ...

...human kinases remains challenging. Small molecule kinase inhibitors usually target soluble kinase domains...Human kinase domain constructs for automated bacterial expression The misregulation of human kinases has...recently compiled the Human Kinase Domain Constructs Kit, a library of human kinase domain constructs developed...Threonine kinases while YopH (residues 164-468, Plasmid 79749) enhances expression of Tyrosine kinases. All...Nucleic Acids Res. 1996. PubMed PMID: 8692687 Tracking autophagy in yeast with Rosella, a pH-sensitive...a target cell. In addition to nucleotide-based linking strategies, post translational protein fusion would...Arumugaswami lab at Cedars-Sinai Medical Center has kindly constructed and deposited an unpublished set of...