...etc.) (4). This has also been identified with AAV2 (5), AAV6 (6) and AAV5 at excitatory connections (3)....tropism and axonal transport of AAV serotypes 1, 2, 5, 6, 8 and 9 in the mouse brain." PloS one 8.9 (2013...PMID: 24694006. PubMed Central PMCID: PMC4137353. 5. Ciesielska, Agnieszka, et al. "Anterograde axonal... Its Fruit Is Sweet The Future? I knew little about adeno-associated Viral Vectors (AAVs) before starting...I’ll provide some tips that will help you think about how to get started with AAVs. Since my research ...understanding of receptor physiology and the effects of metabotropic and ionotropic influences on overall cell function... the biotechnology firm Biogen have recently collaborated to develop “AAV 3.0”, vectors that will have...

...in Sweden. It can be very time consuming to design 5 guide RNAs (gRNAs) targeting each of the 1000 genes...constellations: Doench/Root (2), Zhang (3), Wu (4), Yusa (5), Wang/Lander/Sabatini (6), and Chari/Mali/Church ...PMID: 26190106. PubMed Central PMCID: PMC4559080. 5. Tzelepis, Konstantinos, et al. "A CRISPR dropout ... to design gRNAs for all the purposes discussed above. Please see videos and text on the website for more...Output: A text file containing full information about the suggested gRNAs to use, including all information... hard to read. It´s suggested that you open the above text files, copy the content of the file, and paste...Additional Resources on the Addgene Blog Learn about CRISPR Screens How to Design Your gRNA Tips for ...

...Plasmids and Chromosomes. Microbiol Spectr. 2014 Oct;2(5). doi: 10.1128/microbiolspec.PLAS-0025-2014. PMID:...concatemers in mammalian cells. J Gene Med. 2009 May;11(5):444-53. doi: 10.1002/jgm.1310. PMID: 19263463. Ribeiro...multicopy plasmid stability. Mol Microbiol. 1998 Sep;29(5):1137-45. doi: 10.1046/j.1365-2958.1998.01012.x. PMID...colonies using one of the detection methods described above can help ensure the isolation of the monomeric version...plasmid DNA production platform incorporating low metabolic burden seed-stock and fed-batch fermentation processes...

...implants have been reported by several groups [4, 5, 6]. These have the obvious advantage of not requiring...PMID: 23580530; PubMed Central PMCID: PMC3769938. 5. Iwai Y, et al. A simple head-mountable LED device...distance so ideal implant placement should be about 0.4-0.5mm above the site of your stimulation. The scatter...scatters a great deal once it enters tissue, placement above the site of stimulation guarantees that the majority...instead). Laser stimulation         In the video above, Channelrhodopsin 2 (Chr2) has been expressed in...These lasers are relatively cheap and will run you about $2-3,000/laser. However, an important caveat is ...output of the laser must be controlled. As discussed above, each opsin variant has slightly different activation...

...ideas, applying for grants, and mentoring. Chapter 5: Wei Leong Chew's lab management style and encouraging...getting a lab up and running, and learn a little bit about what it was like for Wei Leong to work in George...

...antibodies against some AAV types, AAV1, 2, 3 and 5 are thought to be as high as 70% among the human population...therapy 10.11 (2003): 964-976. PubMed PMID: 12756417. 5. Chan, Ken Y., et al. "Engineered AAVs for efficient...Vectors Featured Topic Page Learn about CRISPR-AAV vectors Learn about Addgene AAV Quality Control Process...viral infections. She is particularly passionate about HIV cure research and science communication.     ...

...Science Rapper Tom McFadden Learn about SITN's work at DayCon 2015 5 Reasons to Use Reddit for Science...supporters of science communication. To help you think about ways to effectively communicate your science, we... of programs SITN runs and learn how they think about communicating science. Listen to the interview ...

...below! Additional Resources on the Addgene Blog 5 Science Rap Videos and Their Creator: Tom McFadden...researchers time by quickly relaying key points about an unfamiliar topic or lab process. Along with our...goals, or to answer any questions you might have about Addgene. The page is divided into three main sections...a variety of roles at Addgene.  You’ll hear all about their career paths, future goals, and get some useful...

...Joachim’s lab. New microscopy probes and more. Chapter 5 - mTurquoise II and the creation of improved fluorescent...of our educational resources. Listen to hear all about Professor Goedhart, his lab, and some of the tools...

...dTAG-13 at doses as high as 20 µM (see Figures 3 and 5 in Nabet et al., Nature Chemical Biology for examples...degradation.. Nature Chemical Biology. May 2018. 14(5): 431-441. PubMed PMID: 29581585. This reference ...degrade any protein. We are particularly excited about the dTAG system as we can use it to evaluate the...

...requirement. For instance, for Casɸ-2 the PAM sequence is 5’-TBN-3’ (where B is G, T, or C). The authors demonstrate...new YFP variant, mGold. mGold was found to be up to 5-fold more photostable and as bright to mVenus, the... plasmids you need. If you'd ever like to write about a recent plasmid deposit please sign up here.  Here's...

...other options. Commonly used Cpf1 variants, with a  5’ TTTV PAM, improve targeting in AT-rich loci or genomes...activity. Surprisingly, off-target effects across 5 gRNAs with NGG PAM sequences were actually lower for...Blog Learn about Engineered Cas9 PAM Variants Read about CRISPR Base Editing Learn about Homology Directed...editing specificity. We’re excited to learn more about xCas9 - here’s what we know so far! Why do we ...point mutations, also has a strict editing window about 15 nt upstream of the PAM. Even if there’s a PAM...in mammalian cells, finding xCas9 3.7 displayed about the same editing rate as SpCas9 at an NGG PAM. When...are exciting, there’s a lot we still don’t know about xCas9. “I’m not 100% sure xCas9 is going to be flat...

...nm) BD Horizon™ BB515BD Horizon™ PE-CF594Pe-CyBD™5 PEPE-Cy7 FITC Alexa Fluor 488PerCP-CY5.5 PerCP...example, I will describe a basic panel (Table 2, Figure 5) for a hypothetical experiment in the lab. Let's say...FITC channel CD45 BV421   Figure 5: Emission spectra of the fluorophores used in our ...spectrum is located in a range of wavelengths of about 380–700 nm. Photons of lower wavelengths are higher... 650 nm, APC-Cy7's global excitation maximum is about 750 nm. If we were to use a laser that excites at...optimal excitation happens at higher wavelengths of about 750 nm. Source: https://www.bdbiosciences.com/en-us... we start building your first panel, we'll talk about one last principle of color emission. As you can...

...surgeries for viral injections can be found here [5, 6]. Opsin transgenic mice Transgenic mouse lines ...10.1038/nmeth.1808. PMID: 22179551; PMCID: PMC4165888. 5. Harris JA, et al. Adeno-associated viral vectors ...activation and silencing. Nature neuroscience. 2012;15(5):793-802. doi: 10.1038/nn.3078.  PMID: 22446880; PMCID... electrophysiology data too but that’s what collaborators are for). For example, if you study neural pathways...intended brain region and/or cell type (I’ll talk more about this in detail later). But what are the different...of your opsin, but keep you eyes open for posts about the next steps in the coming weeks. Expression of...Addgene. See the Deisseroth lab homepage for details about types of constructs available, instructions for ...

...Additional Resources Quick Guide to All Things Lentivirus 5 Tips for Troubleshooting Viral Transductions Getting...Addgene.org Check out our Viral Service Learn more about our Viral Production Process Find other Viral Vectors...

...being receptive to those who report to you. Chapter 5: The types of science that Crystal works on at Novartis...therapeutics at Novartis - listen to learn more about Crystal’s exciting career path, to gain insight ...Chapter 2: What's a lab group in industry? The collaborative nature of working in industry and how delegating...metrics for success. The importance of thinking about the big picture and not just the next experiment...resources in our Mentoring for Scientists Series Learn about Turning Your PhD into a Career with Joanne Kamens...

... to limitations in light penetration mentioned above (5). Thick static samples Ex: Fixed tissue sections...providing fast imaging along with optical sectioning (5). One such example is the lattice lightsheet that ... Enzymol 361, 1-33 (2003). PubMed PMID: 12624904. 5. Power RM, Huisken J. A guide to light-sheet fluorescence...call Epi-fluorescence microscopes. (See left image above for an example image). Point Scanning Confocal Microscopy... plane to reach the detector and all light from above or below is excluded. Parallelized Confocal Microscopy...sectioning. Point scanning confocal microscopes (see above) often provide the highest quality images for this...

...contaminant we mixed into the original AAV sample at 5%. Detecting 5% contamination is a good start, but to determine...viral genome sequencing (VGS). Why should you care about viral quality control? AAV vectors are produced ... let us know if you have any specific questions about viral DNA NGS you’d like us to discuss. We are also...or at [email protected] to answer your questions about our quality control process! The images in this...

... the desired gRNA target sequence, as well as the 5’ and 3’ homology arm repair template (Fig. 1, Step...of the same type will produce the same unique SapI 5’ overhang, different combinations of connector and...tagged gene. Most insertions can only be detected by labor-intensive processes like PCR or by evaluating visual...repair after a CRISPR/Cas9 induced break. Learn more about these constructs and the cell lines they’ve been...Additional Resources on the Addgene Blog Learn about Fluorescent Protein Timers Check out Photoactivatable...

...Money with the Baby Gel Box Find 5 Smart Phone Apps for Lab Life Learn About the Evolution of Lab Techniques...Digests Make Your Agarose Gel Other Resources Read about Gel Stains on the BitesizeBio Blog ...