We narrowed to 860 results for: PLE

...ccdB aspect, Gateway takes advantage of the same principle that cells will not propagate while expressing...system contains ccdB. A successful insertion will completely replace ccdB with the investigator’s insert of...interest.   CcdB resistant E. coli strains complete the system But ccdB alone isn’t the only key to...most appropriate method for your own experiments. Please check out Addgene’s Choosing a Molecular Cloning...protein involves poisoning of DNA-topoisomerase II complexes.” J Mol Biol. 1992 Aug 5;226(3):735-45. PubMed...

...Assembly, Golden Gate etc.), and try to find the simplest way to do it (i.e. avoid assembling too many fragments...methylation like XbaI, ClaI, etc. If you are doing multiple digestion be sure that the buffers and temperature...Remember to quantify the DNA concentration of all the samples by gel or by nanodrop to assemble the  fragments...molar ratio. Assembly of the fragments There are multiple ways you can assemble the different parts of a...plasmid. Many vectors contain sequences that are complementary to standard sequencing primers. If not, design...

...is guided by an RNA molecule, the gRNA, to a complementary site in the genome where it will proceed to ...degradation. Cas3 DNA digestion can proceed up to multiple kilobases, but only in one direction. Cas3 is ...that Cas14 also indiscriminately cleaves non-complementary ssDNA when engaged by its target sequence. Why...used in lieu of siRNA or shRNA for efficient, multiplexable RNA knockdown. Cas13 has also stirred up interest... the other Cas family members make for useful implements in the scientist toolbox. If your genome engineering...

...conduct research that fails less and helps more people, we need to take sex into account. Gendered Innovations...s Health Initiative Hormone Therapy Trial, for example, found that the study saved lives. Reduced use ...analysis and 2) provides case studies with concrete examples of how gender analysis leads to discovery and ...experiments—which can lead to failed research. For example, an international research team from Norway and...studies that draw from a body of research that is incomplete (because sex was not analyzed) may harm humans...

...extended to include a primer binding site (PBS) complementary to the genome and a reverse transcriptase (RT... the experiment. Now, onto the tips! General principles pegRNA design If you’re just starting out with... nicking the newly synthesized strand before heteroduplex resolution, which can lead to indels (Anzalone...edited strand as a template during DNA repair. Test multiple nick sites, starting with sites about 50 bp upstream...Nureki, O. (2014). Crystal Structure of Cas9 in Complex with Guide RNA and Target DNA. Cell, 156(5), 935...

...all essential embryonic genes are individually depleted via RNA interference (RNAi). Scientists monitor...AID system discovered in plants to conditionally deplete proteins of interest in C. elegans (Zhang et al...Schwartz and Jorgensen, 2018).  These are just some examples of worm resources and tools openly shared by the...Hammarlund, Marc, et al. "The CeNGEN Project: The Complete Gene Expression Map of an Entire Nervous System... system enables versatile conditional protein depletion in C. elegans." Development 142.24 (2015): 4374...

... the xanthine produced via complete demethylation of caffeine. For example, the Barrick lab was able to...ever like to write about a recent plasmid deposit please sign up here.     Listen to the Hot Plasmids episode...living biosensor for measuring caffeine content in samples. The Barrick lab and the UT Austin undergraduate... to grow this strain in minimal media supplemented with various caffeinated beverages, but not with caffeine-free...

... the mutation or replacing the gene. A notable example is the treatment of Severe Combined Immune Deficiency...normally functioning T cell repertoire (1). In this example, it was possible to transduce the T-cells outside...based delivery techniques may hold the key. In the simplest form, translating gene therapies from conceptual...already have ways to prevent AAV infection. For example, prevalence of neutralizing antibodies against ...: 14566358.  2. Hirsch, M. L., et al. "Self-complementary AAV mediates gene targeting and enhances endonuclease...

...CRISPR is a simple and versatile tool for genome engineering, but its utility is dependent on its ability...tiny DNA cutting ninjas that deliver CRISPR to multiple types of targets: primary cells, embryos, and ...helps form DNA and retroviral-particle or VLPs complexes. The team found that "all-in-one" Nanoblades facilitated...their CRISPR cargo into the zygote. As a proof-of-principle, the team used Nanoblades to disrupt the tyrosinase...no detectable editing. Additionally, off-target spleen editing was not detected and Nanoblade injection...

...coordination to more complex situations such as learning and memory or emotional states, for example anxiety or ...and cause variability in the testing group. For example, if an experimenter picks up a mouse from the home...kHz, produced from fluorescent light bulbs, for example) that are outside the humans’ audible spectrum.... have preferences for specific cell types. For example, AAV1 and AAV9 serotypes have strong neuronal tropism...

...ever like to write about a recent plasmid deposit please sign up here.  Here's what you'll find in this ...expanding our ability to study and understand the complex interworking of the 86 billion neurons in the human...Using genetically encoded fluorescent proteins coupled with Fibronectin intrabodies generated with mRNA...speed across these different cell types. In one example, H2B-FT was expressed in murine B cells (BaF3),... the plasmids at Addgene The Cas Hybrid for Multiplexed Editing and screening Applications (CHyMErA) ...

...bit of advice I can suggest, is to give yourself plenty of time when writing a fellowship proposal. Yes... you learn new techniques and interact with new people. If you are looking to move to a different university...there is an increasing realization that, if we want people to keep contributing to science, it would be helpful...audience. My lay paragraph typically includes a couple of sentences of introduction to my area, a line...great information on ELISA troubleshooting and multiplex assays.   Additional Resources on the Addgene ...

...challenges and opportunities for us all.  For example, implementation and compliance for the FLSA changes, including...turn. The first is the passing and upcoming implementation of the Federal Labor Standards Act (FLSA) which...remain mixed in their opinions on the subject – for example, Yale graduate students filed for union elections...challenging, but will be aided by the development and implementation of new strategies designed to cope with current...

...titers measure the amount of viral particles in a sample and are usually based on the presence of a viral...however, is that a p24 ELISA measures all p24 in the sample regardless of whether or not it is incorporated...measures infectious viral particles. One of the si mplest methods of measuring functional titer uses FACS...integrated into its genome and a cell that has had multiple integration events. In addition, promoter choice... cell’s genome. This approach can discriminate multiple integration events, reducing the possibility of...

... ONE-GO biosensors kit By Mike Lacy G protein-coupled receptors (GPCRs) are critical components of many...detection of activation triggered by endogenous GPCR. Example applications include large-scale pharmacological... your research! If they do (or if they don't!), please consider sharing your experience with other researchers...recombinant antibody in human postmortem brain samples. Results show positive staining in tissues from... Multiple Authors...

...specific type of virus you are trying to make. For example, bacteriophages will need to be produced in bacteria...cells. However, some viruses can be produced in multiple different cell types, including plants, mammals...supernatant is typically collected over the course of a couple days to increase yield. The collection process ...preps, lentiviral titration requires an additional couple of days, as it requires transduction of more cells...titrated following purification using qPCR or digital droplet PCR. Insect cells Insect cells are a less common...

...I knew when I first became a manager. Know your people Each person is unique and should be managed accordingly...accordingly.  I wish I knew that when I was “managing” a couple of undergrads back in grad school. Both were hard-working...Myers-Briggs test:  my undergrads were different people! More specifically, they had different learning... saying “I’ve been doing some of X’s work for a couple of weeks now”. I had no idea this was happening... and maybe even the slight awkwardness can spur people to speak up. Once aware of the problem, I was able...

...and look at them under a microscope, fractionate samples to isolate particular organelles and their contents...fluorescent proteins have usurped old methods or complemented them in ways that make them much easier. A special...localized to a specific region of the cell. For example, if the ratio of red/green fluorescence falls within...experimental setup less cumbersome. 3. Portability across multiple systems Terskikh et al. showed that their versatile...in gene expression. In a developing embryo, for example, a researcher could determine whether or not newly...

...realized how perfectly flat the complex is on this surface”….where the complex is docked on the membrane of...like “Oh,…I didn’t quite appreciate that….” For example, when Tom Steitz was presented with a physical ... close to its reported cleavage site on the complementary DNA strand. And this is why we love working ...double-stranded DNA, recognize a PAM sequence, allow the complementary base pairing between the guide RNA and the target...

...criticism, and ask thought-provoking questions. For example, if you’re a virologist presenting at the American...like to say to help build your presentation. For example, when thinking about discussing a figure, jot down...practice, practice: Rehearse your presentation with people inside and outside your field. Your colleagues,...brainstorm potential questions you may receive. People often ask about the relationship of your work to...guilty of this, and have many times answered a completely different question than the intended one. Take...