... package stays in transit for a certain number of days and tracking average transit time for each courier...processes, and the distances the packages needed to travel meant the accessibility varied country to country...courier we contracted with so we can always offer the fastest shipping. However, there were issues that ...Protocols Agarose Gel Electrophoresis Bacterial Transformation How to Design a Primer Creating Bacterial Glycerol...Culture Collections Lentiviral Plasmids Tetracyline Inducible Plasmids Empty Backbones Bacterial ...

...the initial data-generating lab has efficiently trawled their own data? A distinct (perhaps similarly impoverished.... For example, mouse behavioral geneticists may track the motion of multiple animals on different occasions... available, but there remains concern about the extra work this may entail.  So if access to data is important...responsible for storage.  If datasets are too large to transfer to a journal repository, they are likely too large...repositories emerge that allow efficient searching and extraction of information. Looking a little further ahead...albeit unintentional, tendency to work against transparency. This opens science to charges of elitism and...opportunities for research as well as increasing transparency. Thank you to our guest blogger! Jim Woodgett...

...background on the different parameters that your gating strategy will be based upon. We’ll try to keep it short... promise! FSC and SSC: scattering To digitally extract your cells based on their size, the cytometer provides...background. Let’s dive into the praxis of gating strategies. As an example, we’ll use one of my previous ... from a C57BL/6 mouse and analyzed the tumor-infiltrating lymphocytes of that tumor. In Figure 3 you can...Figure 2), single cell events will show up in a straight line one the plot, while duplicates deviate from...Q1). If I wish to employ a more refined gating strategy, I can, of course, also draw multiple individual...Reading a Flow Plot Antibodies 101: Conventional vs. Spectral Flow Cytometry...

.... Plasmids 101: Transformation, Transduction, Bacterial Conjugation, and Transfection by Alyssa Cecchetelli...Plasmid Cloning by Jennifer Tsang There are many strategies for finding your clones in an experiment. Read...Goedhart Guest blogger Joachim Goedhart shares some strategies for generating GFP fusion proteins. From considering...editing. Using a fusion between Cas9 and reverse transcriptase and a prime editing guide RNA, it can introduce...

...Training as a scientist in the academic system has many pluses. I delighted in my graduate school years...nature of academia often means that scientists-in-training rarely get the opportunity to learn about the ...science. I find it ironic that we do all of our training as scientists (5-12 years worth!) with academic... one of the best things you can do during your training is meet interesting people doing interesting things...connection that many people honor. Talking with strangers is usually challenging but does get easier with...you might spend a whole day having to talk with strangers. Be bold in asking all kinds of people to connect...

...after a holiday break filled with family, food, and travel, and you can’t remember where you put your flow...four white binders. Thirty minutes later you’re frustrated and no closer to starting your experiments. Get... deadline again. Organize you experiments, keep track of upcoming grant deadlines, and sync everything...Though electronic lab notebooks (ELNs) have gained traction with pharmaceutical companies, academic labs have...allow you to control who can edit the documents and track the revision history. Now you’ll be able to make... knowing the information is backed up and edit tracking will help you identify when others have added ...

...proteins, or transcriptional regulators. In this system, Cas9 generates a double-stranded DNA break at...are complementary to either side of the double-stranded break in the genomic loci of interest. As the ...groundwater. Now, they’ve recently published data demonstrating that CasX is active in both E. coli and human... Find the CasX plasmids Drug inducible transcriptional activation and genome editing Yu Wang's lab...developed drug inducible CRISPR/Cas9 systems for transcriptional activation and genome editing by fusing Cas9...

... aim of improving professional development and training in a globalized world where individuals can exchange... steps. For example, how to get a fellowship to travel abroad, how to write cover letters, and has suggested... am not sure that I want to pursue the academic track.” How does an International Mentorship Program work...throughout the entire year. In addition to their direct training role as mentors to Spanish mentees, mentors also...University of Valencia, Spain) as the most improved trajectory during the pilot, and Joaquín López (MIT, USA...models based on integrative and eco-social health strategy, and global health equity. She is also Co-founder...its International Mentoring Program Director of Strategic Initiatives. Learn more at https://www.linkedin.com...

...sequences to sequences in a database ExPASy translation tool: Translates a DNA sequence using all six open reading... looking at plasmid maps and viewing sequencing traces, while the paid version provides more tools for...Annotates features, BLASTs, aligns, digests, and translates a sequence.   NEBCutter2: For finding restriction...information, such as expression patterns, post-translational modifications and polymorphisms Codon optimization...your lab protocols using apps to save you time and track your experiments  Resources on Addgene.org: Read...

...chatted about social media analytics and summer travel plans while eating lunch. There was something so...from that, suggest changes to our social media strategy. We ended up implementing some of these changes... with my teammates acted as my safety net and trampoline all at once. I was able to do something that ...of using analytics to make decisions on content strategy. I really believe that the culture at Addgene ...cultivates trust between coworkers and that trust then translated into this incredible experience.  Of course, ...loads of different best practices, create an email strategy, and make the occasional TikTok. I was also responsible...

...19805086. PubMed Central PMCID: PMC2764931. Portugues, Ruben, et al. "Optogenetics in a transparent animal: circuit...of neuroscience. The larvae of zebrafish are translucent and allow for noninvasive live imaging with fluorescent...as spontaneous swimming and escape reflex. These traits, coupled with short generation times and high fecundity... to identify new neurons that trigger the CPG (central pattern generator) (Wyart et al., 2009). The CPG...461.7262 (2009): 407. PubMed PMID: 19759620. PubMed Central PMCID: PMC2770190.  Additional resources on the...

...positively transfected. Alternatively, you can use a selection marker to select transfected cells (for...nickase doesn't have the ability to induce double-stranded breaks like the wildtype Cas9 does. There are ... The human U6 promoter prefers a 'G' at the transcription start site to have high expression, so adding...Before starting your experiment, consider co-transfecting with GFP. This allows you to sort for GFP-positive...resistance cassette, such as PX459). After you co-transfect the CRISPR/Cas system with your homologous recombination...

...company.  Ultragenyx’s goal is to develop novel products for the treatment of rare and ultra-rare diseases...deposited the ClonTracer Barcoding Library, a library that “enables the high-resolution tracking of more than...Dimension Therapeutics, which was acquired by Ultragenyx Pharmaceutical in 2017. As one of the original...the targets that have the biggest potential to transform patients' lives. Previously Stegmeier was the ...

...Wanisch  Listen to the optogenetics podcast segment LOVTRAP is an optogenetic system that was recently developed...modifying EB1. EB1 normally sits on microtubules to attract other proteins involved in their growth and extension...promoting domain. Each half was fused with one of the LOVTRAP domains. When both halves of the modified EB1 protein...separation of its synergistic domains. This paper demonstrated that the migration of a cancer cell line equipped...fluorescent proteins (FP) with improved tissue penetrance and live cell imaging capabilities. Upon developing...Quantification of editing efficiencies simply involves tracking the ratio of mCherry-positive to eGFP positive...

...create a toxic product. The E. coli strain Most E. coli strains can be used to propagate plasmids, but...plasmids from the same group should not be co-transformed, so if you need two plasmids for an experiment...groups. Note the A -C compatibility grouping is an arbitrary designation, and plasmids from the same incompatibility...incompatibility group should not be co-transformed. +Actual copy number varies. See below for additional...stocks, agar stabs, or liquid cultures. Freshly transformed cells also give a higher plasmid yield that colonies...

...investigator. In basic science (as opposed to translational, biomedical, or applied sciences), people conduct...don’t tell us everything there is to know is a strange and exciting revelation. Science in any field can...postdoc) Getting more scientific and professional training after earning a doctorate Considered a "stepping...1-2 years) Often a gap-year position taken to transition to the next career stage (ex: grad school, med...duties ex: order materials, lab maintenance Administrative assistant: assists PI with admin duties Staff...reach out to the PI to get involved. It not only demonstrates initiative and drive, but also makes for an ...that you have new interests and that you want to travel in new directions. During my grad school interviews...

...large inserts can make lentivirus production frustrating - a researcher can complete an entire lentiviral...editing experiments. We aim to alleviate this frustration by producing, titering, testing, and providing...troubleshooting reagent production, because that's a distraction from your primary goal. Getting these quality-controlled...ready-to-use lentivirus from Addgene, you’ll also receive transfer plasmid DNA that you can use along with lentiviral...Addgene Blog Get the Most from Your Lentiviral Transdution Using Virus in Your Research: A Primer for Beginners...Beginners 5 Tips for Troubleshooting Viral Transductions Additional Resources on Addgene.org Visit the...

...across the 11 candidate transcript targets analyzed. CRISPR-Csm presents an attractive alternative for nuclear-RNA...off-target costs. Additionally, this system also demonstrated the ability to be used as an endogenous RNA ...! Colognori D, Trinidad M, Doudna JA. Precise transcript targeting by CRISPR-Csm complexes. Nat Biotechnol...delivering them to a tumor - they used focused ultrasound (FUS) to heat the tumor area, as the image below...activity of CAR-T cells within tumours via focused ultrasound. Nat Biomed Eng. 2021 Nov;5(11):1336-1347. doi...

...responses are varied; one outcome is directed transcription from a given promoter. To harness the power ...the RR (OmpR) via phosphorylation, promoting transcription (see figure below). Subsequent exposure to red...response regulator via phosphorylation, promoting transcription of a GFP reporter. Subsequent exposure to red...system less bulky and easier to use with other strains of E. coli, Schmidl et al. condensed the system...types of proteins, including Cas9 and dCas9-transcription factors, for which low expression levels are... optimal. These tools are compatible with many strains of E. coli, as well as with other plasmid-based...

...disease. CRISPR has made genomic SDM relatively straightforward, but plasmid-based SDM has lagged behind. While... and the nonmethylated PCR products are then transformed into E. coli to undergo nick repair. This method...homologous recombination. In most recombineering strategies, only 3-4 phage proteins are needed and these...containing the desired mutations, to amplify both strands of a plasmid template. In addition to containing...template plasmid, these PCR products are directly transformed into bacteria expressing recombineering proteins...Sci Rep 6:19121. PubMed PMID: 26750263. PubMed Central PMCID: PMC4707547. Additional Resources on the ...