...ask for an amplification protocol with your deposit. This is because repeated amplifications of pooled ... an amplification protocol developed for each individual pooled library; generalized amplification protocols...Plasmids Variable Region Size DNA per Amplification Amplification Type Link to Protocol CRISPR Knockout... this blog, we’ll discuss how to develop an amplification protocol for your pooled library deposit. When...to amplify the sample for further downstream applications. As individual libraries can vary significantly... to consider when creating a pooled library amplification protocol. Created with BioRender.com.   ...Libraries Protocol.   Selecting your strain for amplification Since pooled libraries often contain thousands...

... was founded in early 2018 with the goal of communicating science to non-scientists. Chinmaya Sadangi,...questions about The Addictive Brain and science communication for general audiences. How does The Addictive...Addictive Brain address science communication? At The Addictive Brain, we collaborate with scientists from around...sci-art.  What are your top tips for science communication? It is important to identify your audience ...on the Addgene blog Find all of our science communication posts Sign up to receive our science career ...guide Learn about different careers in science communication Resources on Addgene.org Learn how Addgene...

...researchers involved. Communication is a major challenge when researchers are communicating across disciplines...closely replicates how you’d interact with team members in person, and improves overall communication.  Maintain...Furthermore, this casual communication reflects the type of informal communication you’d likely have with...But these challenges can be exaggerated when communicating across time zones, digitally, and asynchronously...to keep everyone on the same page is to make communication a non-negotiable part of your collaboration....great way to include researchers from different locations. Having virtual face-to-face meetings as opposed... Maintain informal communication In between recurring meetings, you should share interesting results or...

... DNA purification without a kit as well as one way to reuse silica columns from DNA purification kits....consider some ways to purify DNA without a kit. DNA purification kits have advantages: they are convenient and...because you’ve run out of columns. Kit-less DNA purification methods avoid many of the drawbacks of kits ... and then rehydrated. There are some slight modifications depending on the type of DNA you’re purifying...After this, the DNA is ready for use! Plasmid purification and DNA gel extraction with glass syringe filters...Product: plasmids or DNA fragments The plasmid purification and DNA gel extraction method described in Kim...salt (guanidine hydrochloride for the plasmid purification protocol and guanidine isothiocyanate for the...

...the origin of replication/replicon. The replicon is comprised of the origin of replication (ori) and all...   Types of origins of replication There are lots of origins of replication out there so, for simplicity...gene for replication (reference). **F1 is a phage-derived ori that allows for the replication and packaging...control elements. The ori is the place where DNA replication begins, enabling a plasmid to reproduce itself...are generally different from the those used to replicate the host's chromosomal DNA, but they still rely...readily at lower temperatures and allows the replication machinery room to come in and get busy making...derivatives), pSC101, R6K, and 15A. Not all origins of replication are created equal. Some will produce many plasmid...

...previous nosy question! The Application Process Broadly speaking, the PhD application process tends to go something...steps in the application process and usually occurs in the few months leading up to applications opening. ...program. Most applications will recommend that you name two professors in your application that you intend...to continue the application process through a campus visit, a weekend where applications can get to know...strengthen your application and help you decide if a program is a good fit for you. The application process looks...next?” For those finishing up an undergraduate education or ending a Master’s program, the answer might...know that this process only became even more complicated with COVID-19 and standards for hybrid, virtual...

...protein purification. For a more thorough introduction to pCri, please see the original publication cited...expression and purification: a comprehensive review of affinity tags and microbial applications." Biotechnology... Protein purification can be one of the most stressful lab activities. Working with proteins requires...interest can save a lot of time later. Protein purification specialists recommend doing some research, notably...Gomis-Rüth, aims to take the guesswork out of protein purification through simple and efficient multiplexing. pCri...sites should one need to remove the tag post-purification. A sample of pCri constructs: pCri-1a, -4a ...4a and -9a. 6xHis tags enable affinity purification using a Ni-NTA resin. MBP improves protein solubility...

... Here at Addgene, we’re dedicated to advancing and sharing science! In association with the Harvard graduate...chief goals. First, SITN works to bridge the communication gap between scientists and the public and help... SITN helps graduate students develop their communication skills and engage with the public. The organization...students, these opportunities to improve our communication and organizational skills provide great training... science communication...

...plasmids rely on their host cell's replication machinery in order to replicate—but not always to the same extent...described in our previous Origin of Replication post, DNA replication is initiated at the ori and may or...consider the replication mechanism of plasmids under stringent control, such as pSC101. DNA replication and regulation...review how E. coli normally replicate their chromosomal DNA. Chromosomal replication in bacteria is carefully...entire round of replication. Initiation is the primary control point of DNA replication in bacteria, so... the replication forks reach each other on the opposite end of the chromosome, ending replication.   ...chromosomal replication and balance population growth against sustainability. Plasmid DNA replication and regulation...

...Voltage indicators are voltmeters for cells! In this blog we will review how these indicators work, their...fluorescence or voltage indicators.   Viral Vectors and Voltage Indicators As previously mentioned...Enhanced genetically encoded voltage indicators advance their applications in neuroscience. Curr Opin Biomed...Cellular communication and function depend on potential energy. Cellular activities like action potentials...their general applications, and why viruses are essential to their use as biological tools. Why Use Voltage...Voltage Indicators? Electrical signals are integral to many facets of biology. For example, many heart...within a cell are genetically encoded voltage indicators (GEVIs) and chemigenetic sensors. These tools...

...established uses, and intended applications. Within CasPEDIA, there is a classification system, CasID, to help...multiplex ability Experimental guidelines and applications Protein structure and properties Links to ...you want to read literature on the less common applications of an enzyme for your non-traditional experiment...al. (2023). CasPEDIA Database: A Functional Classification System for Class 2 CRISPR-Cas Enzymes [Preprint...I., Iranzo, J., et al. (2019). Evolutionary classification of CRISPR–Cas systems: a burst of class 2 and...

...after application of NE (10 μM) and corresponding pixel-wise ΔF/F0 heatmaps. White insets indicate surface...developing and depositing two new norepinephrine indicators: nLightG and nLightR (Kagiampaki et al., 2023...surface expression of the indicators over white dashed lines. Scale bars, 10 μm (HEK293T), 20 μm (neurons...trialed in two-photon in vivo experiments. nLight indicators are more sensitive to and selective for norepinephrine...They can, however, be studied with the nLight indicators. Speed One of the major advantages of this next...… Patriarchi, T. (2023). Sensitive multicolor indicators for monitoring norepinephrine in vivo. Nature...

...and 3) (5, 3). Benefits of truncated gRNAs Importantly, the use of truncated guides has largely been demonstrated...protospacer, in the genome. Native Cas9 has two catalytic domains, each of which cleaves one strand of DNA... addition to its utility in making targeted modifications to DNA, Cas9 can be reprogrammed to serve as...as a regulator of gene expression. Its catalytic domains can be mutated to inactivate the protein’s nucleolytic...target sites. Regulating gene expression with truncated gRNAs An alternative approach is to modulate Cas9...native Cas9 is complexed with a gRNA that has been truncated such that it exhibits 15 or fewer nucleotides ...should be noted, however, that on rare occasions truncated gRNAs (≤15nt) have been found to retain some ability...

...re more interested in selling a product than communicating your ideas with people or discussing science...to write about in the future. The AMA was also cataloged for permanent reference by the Winnower. The only...I do a lot of public speaking where I enjoy communicating face to face with the audience. I post on LinkedIn...own expertise and your resources for science communication opportunities like an AMA. One of the best things... science communication...

...extension is repeated 25-35 times to exponentially replicate the target DNA of interest. This entire reaction...have been developed for different scientific applications. These PCR methods all use the same basic PCR...the final cycles which would slow or stop PCR amplification. Other times, the PCR reaction may not be 100%...quantified by qPCR. qPCR is used in a variety of applications including gene expression profiling, studying...concentration, unlike qPCR where results can vary across replicates. ddPCR can be used to quantify DNA sequences ...droplet contains the material required for PCR amplification. Following the PCR reactions each droplet is... our blog post “Droplet Digital PCR for AAV Quantification.”   Multiplex PCR Multiplex PCR, as the name...

...xkcd.com Science communication should be fun and engaging for the communicator as well as the audience...many ways options available to today's science communications: podcasts, blogging, videos, lectures, exhibits...in what people think of as "the best way" to communicate their science, meaning that they will heavily...heavily evangelize for the mode of communication they connect best to. And that's really natural - if it works... text or use Facebook. However you choose to communicate your science, some people will love it and others...platforms we use at Addgene: YouTube, for longer educational videos, and Twitter, for sharing updates, new...factors like socioeconomic status and geographic location. For example, not all social media platforms are...

...their use in other clinical applications. In the last decade, viral applications in cancer treatment and ...s unpack the myriad number of emerging viral applications and give you some recommendations for putting...putting viruses to work. Viral applications Viruses work by infecting a host cell (the target of viral infection...expressed. It’s important to note that for all the applications we discuss, there is more than one type of virus...retrovirus, AAV, etc.). When you’ve decided on an application, our Viruses as Biological Tools blog can help...viruses and pick one for your experiment. Research applications Gene expression control Researchers are always...expression control mechanisms require a specific modification of the viral vector for your gene of interest...

...after credible attempts to replicate major findings could not be replicated by other researchers.There...published research is difficult to replicate.  Why is replicating previous research findings so difficult...can be difficult to replicate. Unclear methods Until someone attempts to replicate the results of another...positives in original studies and false negatives in replications are unavoidable, but can be minimized with high...obtaining a different result after conducting a replication study. An important difference between the original... methods, QRPs, and small samples, then when replications do not come out the same as an original research...data, materials, and code Only when you try to replicate the work of other researchers (or your own from...

.... I needed to a) figure out the sources of miscommunication and b) find practical solutions.  My PhD ... for this task. Good scientists learn how to communicate their work differently based on their audience... family is going to vary widely. The way you communicate your science at a poster is going to vary depending...two main problems with Addgene’s cross team communication. Although these real world examples did not ... for the container to be used in cross-team communication A list of “AKA” names, or jargon terms, for ...tracking system. Though you might not have to communicate with a team of software engineers in your lab...experimental protocols. Clarify: Asking for clarification during a meeting can help get everyone on the...

...genetic underpinnings of these processes has been complicated by the lack of precise genetic tools for modulating...recent advancements and offer commentary on their application to neuroscience research. What is CRISPR-based...accessible to neuroscientists, not all neuroscience applications require such optimized tools. This is elegantly...necessitates such molecular gymnastics for sensitive applications such as CRISPR screening. The plasmids used ...used by the Qi Lab for this particular application can be found here.  Why CRISPRa? Although cDNA overexpression...gene activation in select in vitro and in vivo applications, the tools described above make CRISPRa in the...relatively easy and accessible. There are also other applications for which CRISPRa is well-suited, including ...