...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...

...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...

...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...

...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...

...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...

...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...

...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...

...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...

...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...

...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...

...similar for both, though tissue samples do require an extra homogenization step. For this blog, we're going ...but bone or especially fibrous tissues may need extra steps.  Fragmenting  The next step is break the ...your protein of interest is a low abundance transcription factor or cofactor. In such cases, the gentler...Proteinase K to purify the DNA and then determine concentration.  Protein Capture After crosslinking and sonication... purification kit or with a phenol-chloroform extraction.    Fig. 1: Schematic overview of ChIP..., it may be a good idea to add "patience and an extra-methodical approach" to the top of your protocols...

...not entirely. What truly unites us all is the frustration of an experiment failing because of a small, ..., avoidable mistake. Or worse, when you're trapped in a pattern of small errors that significantly impacts...rushing, no matter how hard I tried to build in extra time before the experiment. What fixed it was not...pipette tips to match your plate layout can help you track samples as you add them. Moving your reagents from...the headphones aren’t actually playing music. Nontraditional cues are an option, too - one of my coworkers...and open to new methods, you’ll be able to find strategies and systems that set you up for success in the...

...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 ...

...community. As a result, I have occasionally pondered translating my research into a startup company. However, ...school and turn it into a biotech company. Transitioning an idea from academia to a startup Mike Koeris... biotech startup, Mike explained, his career transitioned away from the bench and became all about finding...their idea at various levels of complexity and abstraction and convince people with diverse backgrounds ...company may begin to prove the technology. This transition takes around 1 – 2 years. The product is the...into it.” Have any other helpful tips to help transition a project from academia to a biotech startup ...

... a lack of transparency in most review processes. Preprints and their feedback, in contrast, are immediately..., and increase their impact. Open Access and Transparency Another major benefit of preprints is their ...record of changes, critiques or endorsements. In contrast, journal-based peer review is usually a confidential...committees encourage and consider preprints as demonstration of productivity.  Part of an Open Scientific...Preprints can be one tool in a combination of strategies for your scientific communication needs. Going...

...a variety of assay types. Be sure to check the spectral properties of the fluorophores you are considering..., each other, and your target. I love FPbase’s Spectra Viewer tool for comparing different fluorophores... (HRP) and Alkaline Phosphatase (AP). Various substrates for each of these enzymes allow you to detect...Endogenous carbohydrates added to antibodies post-translationally. 4) Modifications made to antibodies, such ...remove the incompatible agents by dialysis or gel filtration.    There is, of course, a lot more to explore...Stephenson W, et al (2017) Simultaneous epitope and transcriptome measurement in single cells. Nat Methods 14:...

... this transient construct retains functional Type IIS sites and will be re-digested. In contrast, formation...into a modular cloning strategy. Sometimes referred to as MoClo, this strategy uses the Type IIS restriction...Type IIS restriction enzymes are unique from "traditional" restriction enzymes in that they cleave outside...containing plasmid such as pX330. This cloning strategy not only makes it easy to create a single gRNA-expressing...