...) and pSerOTS-C1* ( 188537 ) a better choice in place of the popular SepOTSλ ( 68292 ). An important technical...

...the Voytas Golden Gate TALEN Kit and are used in place of pTAL1, 2, 3, or 4. For both plasmids sequence...

...Full Target Site (5' to 3'; half-sites in CAPS, spacer in lowercase) aak1 TAL3208 & TAL3209 TCTTTGACTCCCGGAGAGagctggtgggctccgggCCAGGTTCTGGAGGGGGA...

...- Cas9 Mouse CRISPR ( C lustered R egularly I nterspaced S hort P alindromic R epeats) is a microbial ...

...small molecules, light, temperature, or even intracellular metabolite levels. Different plasmids and, often...

...Brd3R from five separate lentiviral plasmids The acetyllysine reader BRD3R promotes human nuclear reprogramming...

...using various delivery methods including intracerebroventricular (ICV) delivery in newborns, and retro-orbital...

...sequence GENIE Project 172908-AAV1 pGP-AAV-syn-FLEX-Ace2N-4AA-mNeon-ST A122D WPRE AAV-mediated expression ...

...plasmids shipped as agar stabs. Iverson, et al. ACS Synth Bio. 2015. PubMed PMID: 26479688. RAS Pathway...modifiers like DNA methyltransferases or histone deacetylases. Unfortunately, these inhibitors are often non-selective... industry, the bioethanol industry, and the pharmaceutical industry, depend on the yeast, Saccharomyces..., including fermented beer and wine, fuels, pharmaceutical ingredients, and recombinant proteins. In order...

...1,000 active makerspaces around the world. Hackerspaces.org maintains a list of active spaces near you. ... can find a makerspace somewhere near you. The place to start looking for your local space is the Hackerspace...@derekja  Resources Hackerspace Website https://wiki.hackerspaces.org/Hackerspaces OpenWorm Project Website...manifested itself in the creation of Hackerspaces and Makerspaces back in 2007, which function as centres...with the OpenWorm Project One such space is the Victoria Makerspace in Canada, which I belong to. The ...  Funding Makerspaces One of the main challenges when starting a new project at a makerspace is finding...competition. In 2010, he founded makerspace.ca, Victoria’s first hackerspace which now offers members and ...

... take a look at the RSpace ELN.  The connected approach in action: how the RSpace ELN supports reproducibility...import them into RSpace. The kind of informal conversations about research that can take place in Slack normally...repositories. RSpace has been integrated with two of the five: Figshare and Dataverse, as well as the DSpace repository... reproduce an experiment documented in RSpace using the RSpace datasets deposited in Figshare or Dataverse...Rory is founder and CEO of Research Space, which provides the RSpace electronic lab notebook. Rory is based...contributed by Rory Macneil, founder of Research Space. There are many types of electronic lab notebooks...reproducibility RSpace was designed in response to requirements that emerged from intensive engagement with...

...which makes them capable of diffusing in the extracellular space; they also have low immunogenicity and cellular...anterograde tracer and injected at several hundred sites across the entire mouse brain. Traced neurons expressed...it’s less likely to be sequestered in the extracellular space. In addition, rAAV2-retro’s altered amino...particles will diffuse long distances within the extracellular space along an axon, this does not seem to be the...brain tissue is injected with compounds that intracellularly spread away from the site of application and...suggest that there are several parallel mechanisms in place (for review see Maday et al., 2014). It is currently...Goshgarian, 2015). Nevertheless, the use of these tracers over the past few decades was essential to build...

...with alternating spacers and repeats, was the original impetus for studying CRISPR. Spacers: Copies of foreign...sequences are called protospacers. Repeats: Bacterial DNA that appears between each spacer in the CRISPR array...sequence matches the guide sequence. PAM: The “protospacer-adjacent motif” present in the non-target strand ...the CRISPR array and include a spacer and a repeat sequence. The spacer sequence is the RNA guide, like...real mouthful of a phrase: Clustered Regularly Interspaced Short Palindromic Repeats. That contrast of simplicity...composed of identical repeat sequences and variable spacer sequences. The array is transcribed and processed... into crRNAs, each including one repeat and one spacer. In bacteria, these crRNAs are bound by Cas proteins...

...provided that target is upstream (5') of a protospacer adjacent motif (PAM) Cas9 endonuclease binding to... and a 3-base pair sequence known as the Protospacer Adjacent Motif or PAM. In order for dsDNA to be cut...that each direct repeat, combined with its adjacent spacer, ultimately encodes a single crRNA. The direct...NNGRR(N)  Acidaminococcus sp. (AsCpf1) and Lachnospiraceae bacterium (LbCpf1) 5' TTTV AsCpf1 RR variant... and the unique stretches of DNA between them “spacers” (see figure below). After years of research, we...pre-crRNA into mature crRNA and tracrRNA binding. The spacer regions, on the other hand, are the unique, foreign...tracrRNA forms the scaffold portion of a gRNA and the “spacer region” forms the target sequence.   Overview...

...question I have is, how did we get here in the first place? How CRISPR innovation affected software development...think we are currently at in the CRISPR software space. The CRISPR research community hasn't had a chance...review process has been challenged with many closely spaced CRISPR software submissions. With few prior publications... most value? This is only one of many decisions faced by those engaging in CRISPR design, but it is a ...significant hurdle made worse by an ever increasing marketplace. Since 2013 about 11 tools have been added every...given the lack of attention to it, it is an obvious place to focus further development toward. Quality control...main goals of the software and should be the first place you look to determine a tool’s suitability. Functions...

...difference. Traces The Traces tab shows the edited and control, non-edited Sanger traces in the region...excluding the PAM sequence. PAM Sequence - The Protospacer Adjacent Motif (PAM) sequence for the nuclease used...as determined by comparing the edited trace to the control trace. In the ICE algorithm, potential editing...and Sanger sequenced. ICE compares these sequence traces to give a detailed analysis of CRISPR editing. ...three tabs - Contributions, Indel Distribution and Traces - provides particular details about the indel profile...the reference sequence derived from the control trace file. On the plot, the green (edited sample) and...calls from the .ab1 file are also shown above each trace. The horizontal black underlined region represents...

...Gairik Sachdeva. Healthcare consulting is a fast-paced field, requiring people who are willing to quickly... consulting? Healthcare consultants advise pharmaceutical companies, biotech start-ups, health insurance... commercial challenges or questions that clients face imminently. Consulting firms deploy case teams, ...all parts of a company to collaborate whenever it faces an important challenge. The types of projects mentioned...uncertainties and a rapidly evolving healthcare space. The consultants who enjoy their jobs most, get ...high levels of performance, and occasionally being faced with “shifting goal posts” as a part of a case, ...consulting can sometimes be challenging. The fast paced nature of projects often means working long hours...

...staining both cell surface and intracellular proteins in parallel, then stain the surface protein first, ... Figure 1: Cells expressing a particular surface receptor are stained with a fluorescent antibody...antibodies. It’s also particularly useful for intracellular staining where binding of large immune complexes...antibody thus amplifying the signal.       Intracellular protein targets, measured by fixing and permeabilizing...permeabilizing membranes For intracellular protein targets, users must first fix proteins within the cells...with formaldehyde first to freeze everything in place followed by alcohol permeabilization. Users should..., fix and permeabilize, and then stain the intracellular target. When using this approach, avoid fixation-sensitive...

...photo of an iodixanol gradient after ultracentrifugation. The interfaces between layers are muddled due to...the use of alt text to describe images in online spaces. Alt text is used by screen readers to provide ...from AAV Purification by Iodixanol Gradient Ultracentrifugation First, you’ll want to identify the reader...for AAV Purification by Iodixanol Gradient Ultracentrifugation. The reader may be, for example, a graduate...between a gradient tube before and after ultracentrifugation so that a scientist can extract virus from...details of collecting the purified AAV–that the interfaces between layers are blurred after centrifugation...the syringe needle is inserted bevel-up at the interface between the 40% and 60% layers. Putting them all...

...antibodies for other sample types and for non-surface (intracellular) antigens. Unfortunately, antibodies frequently... control. This can involve cell surface labeling and/or intracellular labeling (or both). It is important...selectivity for cell surface labeling may become less selective if used in intracellular labeling because ...the intracellular compartment may contain cross-reacting antigens not present on the cell surface. The...best to determine this by testing them. For intracellular targets, we usually test 3 different fix/perms...helpful to try all three when looking to label an intracellular target.   Figure 1: HCT 116 WT and SYT1...optimizing fixation/permeabilization reagents for intracellular targets. Since knockout experiments can be ...