...landing sites,” then the recombinase handles the installation of the donor sequence. The latest work evolves...epigenetic silencer Recombinant antibodies for marking astrocytes MagIC-Cryo-EM for sample enrichment and structure...PRC2 with RvLEAMshort (1:6 molar ratio) with 10 minutes of glutaraldehyde crosslinking: representative ...sequences containing attB/P recombinase landing sites on opposite DNA strands. When the prime editor releases...sequence with the attB/P landing site. A recombinase integrates a sequence from a donor plasmid containing a ...resulting in the integrated sequence between two attR/L sites. Image reused from Pandey et al. 2024 under CC-BY...autoinhibition and stimulating the methylation of nearby CpG sites on the target DNA, and thus inactivating the promoter...

...Addgene. What you might not know is that Addgene distributes curated collections of plasmids as kits with ...pExpression vectors) plasmids. The type IIS restriction sites used in these vectors are of low abundance (meaning...yeast, mammalian cells, insects, and even Xenopus oocytes. The expression vectors additionally contain various...protein tags (His, Avi, and GFP) as well as protease sites to facilitate protein purification. We hope this... Using a variety of GFP and RFP fusions as their test genes, the researchers at the BIOFAB showed that...peptides to facilitate protein folding and/or protease sites to remove the epitope tags. Different plasmids in...library of eGFP-tagged human ORF clones to allow testing and comparison of different cell-free expression...

...Design-Build-Test-Learn cycles. If you’re not familiar with this, it’s a method for building and testing new products...that process!   Figure 1: The Design-Build-Test-Learn cycle adapted for video making.   Cycle...rolled several long takes, and we cut it up in post.  Test: Overall, it worked. We pulled it off! Check out...Nonprofit Day, which was the perfect opportunity to test shooting over Zoom, everyone’s new favorite third-party...to some volunteers, who filmed with their phones. Test: I loved the end result, but it was clear this wasn...functions, including starting and stopping recording).  Test: It was unbelievable how well this worked! Interviewees...would remain masked, it freed us up to be creative.  Test: Our first official shoot went off with almost no...

...conventional cloning (to introduce or remove restriction sites), in mapping of regulatory elements (to mutate promoters...parent vector). Also shown are the “B” restriction sites, which flank the targeted region. Upon PCR amplification... restriction site through mutagenesis vastly facilitates the subsequent process of screening for succesfully...maintained. This overlap ensures that the PCR generates a nicked circle rather than a linear product (...Figure B, digest A). In contrast, if your mutation ablates a restriction site, digestion with the appropriate...portion of the plasmid with a primer pair that generates a linear product. The mutated product can then...the original plasmid using the unique restriction sites. Alternatively, if you want to make your protein...

... µL size serotype testing) pAAV-CAG-GFP 50465 AAV8, 9 (20 µL size serotype testing) AAV-hSyn-EGFP ...size serotype testing) pAAV-CAG-GFP 50465 AAV1, 2, 5, rg (20 µL size serotype testing) pAAV-hSyn-EGFP...Our new AAVs include: Control AAVs for serotype testing jGCaMP7c for calcium detection PSAM4 GlyR for chemogenetic...control AAV that scientists can use for serotype testing. These samples are available in various serotypes...encoding GFP. Check out our inventory of serotype testing AAV New control AAVs Plasmid Serotype Name ...are especially useful for studies in non-human primates, where fusion fluorescent proteins can affect ...

...with >3,500 unique isoforms transcribed from these sites. The Zhang lab generated a comprehensive library...TF to determine how its presence influences cell fates. The atlas of directed differentiation To understand...endeavor, a map of possible TF-induced cellular states was generated (Joung et al., 2023). Fig....single cell RNA-seq was performed to map TFs to cell fates. (Adapted from Joung et al.)    This atlas linked...and down-regulated pathways. Individual cellular fates were then matched with candidate TFs responsible...initiate disease onset, providing both a tool to test prophylactics on and a program that any researcher... generate unique cell types. Of the 24 TFs they tested, 22 expressed markers consistent with the cell ...

... event that homology-based searches return no candidates. Our strategy: Express random fragments of the...library worth using if at least 40% of colonies tested showed correctly sized inserts following colony...for single colonies (this usually requires 10-20 plates) (see box below). This initial transformation should...colonies (150-200 per plate) across 20 fresh selective plates. Step 2: Express random phage DNA fragments in...Several protein induction conditions may need to be tested, as with any protein purification, but overnight...protein induction is done, peel the membranes off the plates and place each onto a lysis solution-soaked filter... strongly to nitrocellulose, over-incubation of plates can lead to high background that obscures the desired...

...differences between these approaches, and provide updates on how best to design gRNAs. You can also find ...using S. pyogenes Cas9 (SpCas9), potential target sites are both [5’-20nt-NGG] and [5’-CCN-20nt], as it ...strand of DNA. As a rule of thumb, we avoid target sites that code for amino acids near the N’ terminus of...annotated start codon. Likewise, we avoid target sites that code for amino acids close to the C’ terminus.... For a 1 kilobase gene, since potential target sites occur ~1 in every 8 nucleotides, restricting gRNAs...this can decrease the number of guides one needs to test in the lab. Importantly, for any modification of...has been established, and can be used to identify sites that are likely to give rise to an off-target effect...

...STEM graduates and postdocs in academia, industry and other areas of employment? How many graduates and...related to the structure of the biomedical workforce. Quotes taken directly from participants (summarized in...per PI” “Lack of awareness about how the system operates and functions” “Complete lack of information on...of academic research (4). There are currently 17 sites around the USA undertaking BEST programs, including...regulatory, or clinical environment. It remains contested, however, whether a better solution would be to...grassroots organisations that represent postdocs and institutes that employ them, the diversity of trainee backgrounds... and the many different paths that biomedical graduates take in their future careers. To participate in...

... handle most of these intermediates, with each one acting on specific substrates and in distinct cell ...stranded, while ssRNA viruses can generate dsRNA intermediates as part of the viral replication process. dsDNA...transcription. In an uninfected cell, dsRNA rarely circulates, but upon viral infection, this nucleic acid ...acid can build up. The foreign dsRNA activates the innate immune response machinery of the host. This pathway...virus, you could introduce some of its dsRNA intermediates into a healthy individual to provoke an immune...molecules designed to degrade mRNA transcripts. RNAi activates the RNA-induced silencing complexes (RISCs), which...DNA template, inherently generating RNA:DNA intermediates in the process. These transcription-coupled ...

... genuinely excites them, and reach out to the PI to get involved. It not only demonstrates initiative ...problems. Biomedical research, for instance, contributes heavily to medical practice, and understanding...tasks Principle investigator (PI) Runs the lab Writes a lot of grants Varying degrees of engagement with...helped me cultivate stronger relationships with my labmates. Sometimes what I thought was a dumb and very ...Google is a great resource too.) Connect with your labmates and mentors Many of the grad students and postdocs...resources for career advice and advancement, my labmates have been some of my closest friends and have ...often have scholarships and funds dedicated for undergraduates interested in doing research and attending ...

...ligate many undesirable structures, including substrates containing gaps or DNA base pair mismatches (...such as Taq DNA Ligase, which only reacts with substrates containing no gaps, and will not join any fragments...simultaneous interrogation of multiple potential SNP sites (15). The related LCR technique uses two sets of...generally prefer fully Watson-Crick base-paired dsDNA substrates to those containing one or more mismatches. However...pyrimidine mismatches are typically worse ligation substrates than pyrimidine:purine mismatches. Helix stability...naturally able to discriminate against ligating substrates containing base pair mismatches (i.e., are "higher...reduces the concentration of annealed mismatched substrates (expected to have a slightly lower Tm around ...

...your Eppendorf tubes, hang up your pipettes, and seal up those plates – it’s time to turn your attention...to announce the winners of the 2022 #DeckTheLab contest!  and it's..... Michael Früh submission, which...2022   Congratulations to all our #DeckTheLab contestants!           View this post on Instagram                     ...attention to decorating! To join the contest, you need only decorate your lab, take photos, and then share them... enter: Post your picture or video for either contest on Twitter or Instagram by December 31, 2022.  ...a lab supply tree and set a challenge to my lab mates. They did not disappoint. #deckthelab pic.twitter.com...

... of Antonello Bonci, Director of the National Institutes of Drug Abuse Intramural Research Program (NIH-NIDA... I work in), an obvious behavioral experiment to test for sufficiency of your circuit of interest in promoting..., on the other hand, would not be an appropriate test.  Of course, proper controls are required and include...sutures and subsequent determination of the proper coordinates of your brain region of interest (as shown in...identical for all of these surgeries. Identify coordinates through a rodent brain atlas or research papers...prior to using the virus. You may need to adjust coordinates based on your own method of identifying bregma...readout. The advantage is you can “zero” your coordinates at bregma which allows you to quickly and reliably...

...dye-nitrilotriacetic acid (NTA) conjugates (Kapanidis, Ebright, & Ebright, 2001). These Cy/NTA conjugates work the same ... What are the proper controls needed to pressure-test your FRET assay to make sure it is working? This...fluorophores to the different biomolecules you’d like to test for interaction. In fact, targeting a FRET fluorophore...attached fluorophore. However, since tRNA suppression competes with natural chain termination at stop codons,... only in a few cell types (E. coli and Xenopus oocytes, mainly). Nevertheless, tRNA suppression plasmids...tag from 6 to 10 residues. While these Cy/NTA conjugates are not commercially available, they are easily... the donor and the acceptor fluorophore binding sites are adjacent to each other on the same protein and...

...regularly, and mix samples very well.  Use duplicates or triplicates to increase the accuracy of your experiment...viral vector production facility, food allergy testing lab, and the grad student down the hall from you...-fold dilution series. Each step of the series dilutes by 1:2 for a series that ranges from 1:2–1:32. ...standard dilution series in duplicate or triplicate. Replicates allows you to calculate the curve’s coefficient...replicate measurement divided by the mean of all the replicates multiplied by 100. To calculate the %CV of the...ideal %CV is less than 15%. You should also run replicates for all of the samples in the assay, as this ...these.  Review previous literature or results from labmates to get a sense of the concentrations you might...

... order fulfilment from an average of 40 minutes to 10 minutes. While users can’t tangibly see these improvements...each project. This creates accountability on the part of each developer and creates trust amongst the ...collaborate with the Product Team who in turn collaborates with stakeholders to resolve the questions.  ...would be staying on top of any third party library updates. Third party libraries are reusable software components...with their third party libraries and any security updates. Most of the time, updating these libraries is ...this year, which means there will be no further updates, security or otherwise. This might sound like Greek...

...CNS cell types, including microglia, astrocytes, and oligodendrocytes, provided that reporter genes specific...activators to induce gene transcription at genomic sites targeted by specific sgRNAs. Its RNA-guided nature...packaging limits (i.e., how many sgRNA expression cassettes will fit). Cre-dependent expression of the SPH...selectively target these different transcriptional start sites in order to better understand isoform-specific gene...activity (hence, “immediate-early”) and consequently regulates neuronal physiology by promoting the development...gene possesses 9 distinct transcriptional start sites, all of which yield the same BDNF protein. To demonstrate...cell types (microglia are a notable exception) necessitates such molecular gymnastics for sensitive applications...

...called a Kirby-Bauer disk diffusion test or Kirby-Bauer susceptibility test (Figure 2), and requires at least...shows how to perform a Kirby-Bauer disk diffusion test and check bacteria for antibiotic resistance. Source...bacteria are causing the infection before they can test for resistance. Growing and isolating the infecting...infecting bacteria can take 24 hours or more, and testing for resistance often takes an additional 18-24 hours...available. For routine infections, like strep throat, testing for antibiotic resistance isn’t commonly performed...infections is so rare. When antibiotic resistance testing is requested clinically, this involves growing ... treated. We can prescribe antibiotics for the shortest time period possible to effectively manage a patient...

... bacterial ARG from Serratia sp. 39006. We next tested bARGSer with several inducible expression systems.../L yeast extract, and 5 g/L NaCl) or on LB-agar plates containing glucose and L-arabinose at 37oC for ... arabinose and colonies on arabinose-containing plates will be visibly white, and gas vesicles in cells...from left to right: colony whiteness on inducer plates, white inclusions in cells with GVs via phase contrast...mARGs also had a large genetic footprint (two cassettes + a booster cassette), their expression heterogeneity...mARGs come as a set of two PiggyBac integratable cassettes, both with doxycycline inducible promoters (Fig...increased ultrasound contrast. In all the cell lines we tested, 1 µg/mL doxycycline was sufficient to saturate...