...Your time is precious. Here are a few things to consider before you commit: Double check to make sure ...can be a particularly good resource if you are considering using a reagent for “off-label” purposes. When...about using Google for serious science questions, consider using Google Scholar, which searches scientific...email you’re writing is probably not directly responsible for the problem. We’re here to help. You've ... few days without getting a response, however, consider following up. Ask for a timeline if you are ...upfront how long it will take to get an answer, consider asking to avoid unrealistic expectations. And...

...details on how they work!) Figure 1: Base transitions are edits between the purine bases (adenine, ...limitation to this approach. CBEs and ABEs are base transition editors. They adjust the side groups of DNA bases...task, not a job for a single enzyme. Base transitions Base transversions C → TG → A A → GT → C ...some cases, the same repair pathways that were considered problematic in the development of CBEs and ABEs.... Like other ABEs, the editor they were using consisted of Cas9 fused to an engineered adenosine deaminase...rapidly. If it follows a similar path to base transition editing, we can expect several generations of...to repair DSBs. Base Editing terms   Base transition Base changes between purines (A ↔ G) or between...

...infects host cells through binding the human angiotensin converting enzyme 2 (ACE2) cell-surface receptor...receptor. The spike protein RBD responsible for binding ACE2 is the target of most anti-SARS-CoV-2 antibodies..., homogeneous time resolved fluorescence, and suspension bead assay (Figure 2). After finding no cross-reactivity...scFVs to cognate aaRS antigens as measured by suspension bead assay. Image from  Preger et al., 2020. ... a compact Cas9 ortholog from Staphylococcus lugdunensis that recognizes a NNGG PAM and has comparable...expressing GCaMP6f with this promoter. Image calcium transients in PV interneurons with AAV-S5E2-GCaMP6f are ...

...solution against a part of the body very sensitive to heat- the inside of the wrist or the cheek. (Parents ...work with, require extra serum to survive at low densities - adding to the expense of dilution cloning. ... transfected cells into 3 plates at different densities for selection). A 2% agarose solution is then ...plates containing selection agent at different densities, then dispose of the plates that do not appear...such as the Viaflo from Integra Biosciences) to transition from a 48-well cell plate to a 96-well assay ...

...class of FPs, the FP timers, add an entire new dimension to monitoring localization; using FP timers, researchers...production of dsRed E5. This timer predictably transitions from green to red fluorescence (500 to 580 nm...However, this initial timer was a tetramer with a propensity to aggregate in cells (Tsuboi et al.), limiting... in Xenopus embryos. Of course, timers may be sensitive to changes in oxygen, temperature, and pH, but...et al. developed monomeric FP timers with less propensity to aggregate. These monomeric FP timers (mK-GO... that differentiates human IPSCs into glucose-sensitive insulin-secreting beta-like cells." Nature communications...

...is its simplicity and versatility. CRISPR/Cas9 consists of two components: the non-specific endonuclease...libraries (referred to simply as “CRISPR libraries”) consist of a heterogeneous population of gRNA-containing... is right for you There are several factors to consider when selecting a CRISPR library for your experiments...identified using next-generation sequencing (E). Considerations and tips for successful screens Next-generation...check out Addgene’s 2016 publication “Practical Considerations for Using Pooled Lentiviral CRISPR Libraries...Waxmonsky NC, Swanson LE, Fan M (2016) Practical Considerations for Using Pooled Lentiviral CRISPR Libraries...

...As a person with many roles and responsibilities, I have to think a lot about how to balance those roles...meeting and a means to request funding for events, consider reaching out to it. PTAs have more flexibility...schooler himself and b) I had the background to be considered an authority in this area. What is genome engineering...presentation, she said “it was such a valuable extension to our curriculum and offered engaging examples...Educational Progress (NAEP), virtually none were considered proficient in science, regardless of race or ...posts Resources on Addgene.org Check out our comprehensive CRISPR guide Find CRISPR plasmids and resources...

... blog post discusses some of the key things to consider when performing colony PCR.   Designing colony...for the presence of many different inserts. The downside: this type of primer does not provide information...information about insert orientation, then you might consider designing orientation-specific primers. Blunt ...day, you can save the leftover bacteria-water suspension and use them to start cultures of your positive...up PCR reactions: The remaining bacteria-water suspension will serve as the template for your PCR reaction...tool to have in your molecular biology tool kit. Consider giving it a try next time you’re screening for...

...provenance (this is the history of the cell line consisting of records of the origin and propagation of the...cell line contamination: Epithelial Mesenchymal Transition Is Required for Acquisition of Anoikis Resistance... The same test is so good that it is used as a forensic technique (aka DNA fingerprinting) that can place...convince people of some of the basic tenets of responsible cell culture outlined in this article. Now many...studied biochemistry at Warwick University, gaining extensive cell culture research experience through working...for a range of testing, from animal doping to forensics, DNA sequencing to drug testing. You can find ...

...cells, thus expressing additional EBNA-1 transiently. The transiently expressed EBNA-1 increases protein expression...field [4]. Three major reprogramming methods are considered when generating hiPSCs for clinical purposes:...purposes: mRNA transfection- this allows transient expression of the reprogramming factors Sendai virus- a non-integrating...efficiently [5]. However, the Sendai virus method is expensive, especially if used to make clinical grade hiPSCs...OCT3/4 and shRNA against p53 pCXWB-EBNA1 37624 transient EBNA-1   Many thanks to our guest blogger,...

... elaborated on later. As for what job fairs to consider, some of the best options are the Naturejobs Career...most relevant experience at the top. You may also consider visiting any of the employer’s career pages to... apply for the position. This post on Business Insider nicely summarizes good job fair practice. Career...’t have a particular employer in mind, you may consider starting with indeed.com or monster.com, where... important to realize that you have to take responsibility for your own professional development, not ... Reading and Outside Resources http://www.businessinsider.com/how-to-get-the-most-out-of-job-fairs-2014...

...includes the aforementioned internal control and is consistent with the result in Figure 1 could look like this...associated with the independent variable should be consistent over time and from different preparations of ...replicate control is important, it speaks to the consistency of the reagents and hands performing the experiment...experiment, not the reproducibility or consistency of the observed phenotype associated with the independent...replicates control for the reproducibility and consistency of the observed phenotype associated with the...may be, proper controls are a basic tenet of responsible scientific inquiry and investigation. References...

...express our gene of interest. This element is responsible for initiating the transcription of your insert...eukaryotes have muliple polymerases which are each responsible for a specific subset of RNAs. To gain this specificity...transcript     Description Expression Additional considerations CMV General expression mRNA Strong mammalian...elongation factor 1 alpha Constitutive  Tends to give consistent expression regardless of cell type or physiology... used for Description Expression Additional considerations T7 in vitro transcription/ general expression...regulatable  Often paired with the temperature sensitive cI857 repressor. T3 in vitro transcription/general...

...these particularly energy-intensive systems in a more environmentally responsible manner.  Other ways to ...service items we employ dozens of pieces of energy-intensive lab equipment on a daily basis. By the beginning...100% renewable electricity! A (not so) shocking transition  In early July the Town of Watertown announced...substantially reduced in an attempt to save energy. The transition to 100% renewable electricity will allow us to...

...synthesized enzyme. Biotin protein ligase (BPL) is responsible for this highly specific post-translational protein...biotinylated. In E.coli for example, the BPL, BirA, is responsible for biotinylating only one lysine in the entire...Esmond E. Snell was able to purify the protein responsible for binding biotin, avidin. Avidin was aptly ...Kresege et al., 2004). Avidin is a glycoprotein that consists of four identical subunits that can each bind ...non-selectively label DNA and RNA. Photobiotin acetate consists of a biotin molecule attached to a photoreactive...Photobiotinylation is sometimes easier and less expensive than other enzymatic methods. Scientists have ...

...Faculty Position. This edition has substantial expansions on the chalk talks and Zoom interviews sections... and mentoring to several people that found my insights helpful. All went on to get faculty positions ...understand your proposal to give you proper consideration. 3. Present an easy to follow seminar See item...Implications for human disease" is vague. Instead, consider "A mechanistic understanding of this system will...free!) contains strategies, hints, anecdotes, and insights. After I wrote the first version of my manual,...publish this manual and earn some cash for my expensive habits of running shoes and single malt whiskies...

...being realized. There are a variety of reasons to consider using genome editing to change the genetic code...sequence and ~75 nucleotide scaffold sequence) is consistent between plants and other organisms, but the promoter...CRISPR components can be expressed stably or transiently depending on the delivery method and cell type...CRISPR components can be delivered and expressed transiently using a standard detergent, Polyethylene Glycol...GOI) into plant cells. A) The system typically consists of a Ti plasmid, into which the GOI is inserted...deliver CRISPR-Cas9 to plant cells and should be considered alternative to PEG or agrobacterium-mediated ...

...change on the opposite strand). Both are base transition editors: converting one purine base to the other...pyrimidine base to the other. Figure 1: Base transition editors catalyze purine-to-purine or pyrimidine-to-pyrimidine...BioRender.com. How does base editing work? Base transition editing requires three elements. They are, broadly... in David Liu’s lab in 2016. History of base transition editing Cytosine base editing The beginnings ...frequency of 1.2%. Although the lab did not conduct comprehensive genome-wide studies of ABE specificity, their... a similar approach. Dali Li’s lab’s A&C-BEmax consists of a fusion of both cytidine and adenosine deaminases...The other 8 base-to-base changes require Base Transition Editors, which will be discussed in another post...

... concern, or nothing to worry about all. Let’s consider several scenarios below for a CRISPR experiment...An off-target rate of 5% is far too high when considering human patients and the diseases associated with...many more. While such software typically only considers DNA sequence and doesn’t take into account chromatin...the genome to reduce off-targets, it is time to consider your Cas enzyme. Even if there are similar sequences...unnecessary for many applications, it is the only comprehensive method available. Targeted sequencing approaches...must be quantified slightly differently. For a comprehensive approach, RNA-seq would be an appropriate method...

...individual neurons, including dendritic and axonal extensions and, most importantly, any connections the neurons...neuroanatomy and neural connectivity, providing valuable insights for neuronal communication. Viral vectors and...synaptic step, thus preventing further widespread expansion of RABV. This improvement resulted in a clearer... Not all measures of transport efficiency are consistent which may be caused by the diversity of techniques...the field of AAV vectors. Conclusion An ideal comprehensive toolset for neuronal tracing and network identification... wheat germ agglutinin (WGA) as a new, highly sensitive retrograde tracer in the rat brain hippocampal...nucleus Rabies and neuronal tracing Important considerations when using AAVs Resources on Addgene.org ...