...Cre-lox or one of the FLP-FRT systems in your experiments. Mutated FRT and loxP sites in Flex Vectors –... – inversion only once All recombination events mediated by FLP or Cre are reversible. Whereas the excision...individual neurons one could reduce the plasmids amounts to reduce transfection efficiency. However, if ...recombinase expression while proteins needed for later experiments (cellular tracking for instance) can be controlled... inducible Cre versions can be used in these experiments. This elegant combination of Cre, FLP, and inducible...

...upregulation of essentially any gene by fusing mutants of Cas9 deficient in DNA cutting activity to a ...cannot help you pass. In addition, activation experiments often require quite a bit of tuning before you...wide variety of activators you can use for your experiments. We have found that SAM (Konermann et al., 2014...be true given the results of previous RNA-seq experiments (Konermann et al., 2014, Chavez et al., 2014,...works reasonably well.  Best of luck on your experiments!  Many thanks to our guest bloggers Marcelle...

...promote homologous recombination of linear DNA fragments. Since recombineering does not contain a selection... in the Cas9 or spacer sequence, and these rare events are easily detectable using PCR. The system also...can be also linearized with XbaI to insert extra elements, like a repair template, using Gibson Assembly...system called INTEGRATE (Insertion of Transposable Elements by Guide RNA-Assisted Targeting) and the Zhang...transposition. INTEGRATE consists of four major components including (1) a CRISPR RNA, (2) four proteins...transcription factors, phage proteins, and RNAP components (Dong et al., 2018). They found a transcription...

...Researchers have designed a variety of photoswitchable elements into many other types of proteins. Here, we’ll...sword Microtubules are one of the most important components of the cytoskeleton. They provide cells with ... will surely be useful for a wide variety of experiments. Opto-RhoGEFs manipulate cell morphology and ...phospholipase D (PLD). Now they present a set of variants called SuperPLDs with 10, 30, 50, and 100 times... and cytotoxicity better than the 50x and 100x variants. But those super-high efficiency constructs are...

...For such a therapy to be successful, multiple requirements must be met. First, CRISPR would need to be ...desirable. Should stem cell editing occur, the CRISPR components would only need to be expressed for a short period...skipping therapies would be applicable to 80% of DMD patients. Figure 1: Exon skipping strategies for...the technology is advancing, precision editing treatments may also be closer than we might expect.   References...expression in cells from Duchenne muscular dystrophy patients through genomic excision of exon 51 by zinc finger...

...CRISPR, Addgene gave to me: Functional Genetic Variants Revealed by Massively Parallel Precise Genome ...prime editing systems by manipulating cellular determinants of editing outcomes🎵 On the 5th day of CRISPR...CRISPR, Addgene gave to me: High-fidelity Cas13 variants for targeted RNA degradation with minimal collateral...prime editor with enhanced editing efficiency in plants 🎵 ...

...consider strategies using other types of affinity reagents like fluorescently-labeled nanobodies or scFvs... label such as horseradish peroxidase (HRP). Experiments using enzymatic labeling can be difficult to ...t forget the controls! The necessary control experiments will vary depending on your specific experimental...design. Like selecting an antibody, some of the requirements are similar to standard IF. Here, we’ll walk...intensity patterns are similar between the two experiments. If not, it might be that the other antibodies...

...PCR-based cervical cancer screens. This third grant represents the continuation of our efforts to provide resources...Bengyella and Kwabena Duedu who were awarded similar grants back in June, Gladys will be using plasmids in ...Seeding Labs Plasmid Grant to a third round of applicants at the start of 2017 and would like to thank ...Superhero 5k Find New Cancer Resources Learn About Our Grants to Rare Disease Research Resources on Addgene....

...community submissions for Addgene and CLOVER AAV experiments - and we've updated our submission process so... are selecting AAVs that will work for their experiments, and a key step for that is sharing the data ... negative data helps others to design better experiments (and wouldn't it be nice if those weeks and months... the Data Hub to help select an AAV for your experiments, let us know ([email protected]) and we might...

...understand how to make new treatments and technologies accessible to patients. And what better place to... through engagement with your local or state governments. Most communities have opportunities to join ...boards, support ballot initiatives, deliver public comments, meet with local representatives, or even run ...policy fellowships in federal, state, and local governments, professional societies, industry, and beyond...

...-stimulate mammalian neurons and alter synaptic events: a new technology was born, and the term “optogenetics...infect defined neuronal subpopulations, and this represents a major downside in optogenetics (Belzung et ... delivery of Cre recombinase in mouse primary afferents." Scientific reports 8.1 (2018): 7321. PubMed ...and Peter Hegemann. "Rhodopsin-regulated calcium currents in Chlamydomonas." Nature 351.6326 (1991): 489...neurons: tools, applications, and experimental constraints." Neuron 95.3 (2017): 504-529. PubMed PMID: 28772120...

... and what makes it difficult for many graduate students. As part of this discussion, we mention graphs...school. This is, of course, not real data, but many students discuss the third year dip (see graph). Niroshi...about picking a post-doc project 19:55 - 22:40 : Improvements that can be made to the grad school process ...'s favorite advice to give to younger graduate students   Check Out Angela Depace's Talk on "Precision...

...key to its efficient use in screening experiments; large amounts of virus are required to ensure that ...plasmid in 2015. In line with all of the exciting developments surrounding CRISPR genome engineering in the... We thank our many depositors for making these reagents available to the research community through Addgene...

...together. Second, KTRs are easy to design. The key determinants of KTR function are the positions of the phosphorylatable...measure kinase activity in specific subcellular compartments. Second, because KTRs shuttle between the nucleus...another variable to consider when designing KTR experiments. Third, KTRs are not well suited for monitoring... Regot, S., & Covert, M.W. (2018). Live-cell measurements of kinase activity in single cells using translocation...

...are 24 variants of the cas14 gene that cluster into 3 subgroups (cas14a-c). All these variants share the... provide bacteria protection against infectious agents. Several enzymes behind this immunity have already...many others are still unknown. Identifying Cas14 variants In this quest to discover new and maybe more efficient...

...able to talk and own these accomplishments. Employers and academic departments want to be able to tell that...Leadership and management Take a lead from the amazing students at Washington University who started the BALSA...like FierceBiotech to “learn the business” Attend events with local networking and trade organizations ...

...used these knockout mice to show exactly which components of the known pathway that were essential for ... rapidly performed by targeting all potential components of a signaling pathway. For example, one could...PD-L1 on a cancer cell). In this screen, known components of the cytokine signaling pathway could be targeted...target surface proteins to determine which contact points are involved in a studied cellular behavior. This...accelerated immensely when we share knowledge and reagents. I have not intentionally excluded any libraries...

...following the trend and encouraging departments to showcase the work of students and staff by holding scientific...method to explore the inner structures of delicate plants and other organisms without destroying the original...organisms on your lab bench? Let us know in the comments.    Additional Resources on the Addgene Blog ...

... is a database that provides information about reagents, funding and volunteer opportunities, protocols... hopefully help guide policies and ultimately treatments in a timely fashion.”  With this in mind, Bourque...including datasets and platforms for discussing new developments, and more. As researchers around the world continue...community work together to share open protocols, reagents, and other initiatives for #COVID19 research. ...

...peak absorption wavelengths and light intensity requirements of different opsins, studies have shown that...generates periodic motor commands for rhythmic movements like spontaneous swimming. To do so, the lab expressed... behaviors, the researchers measured the tail movements of 5 day old fish. The larvae were immobilized... groups of neurons that elicited similar tail movements, they were able to narrow down the possible specific...