...protocols and performing rigorous quality control (QC). The specific QC experiments performed varies for ...of Pennsylvania Vector Core. Transfections are performed using the transfer plasmid, a plasmid encoding...and stored at -80 °C. Titer Titering is either performed by Addgene or by the University of Pennsylvania...Pennsylvania Vector Core. In general, titering is performed by the facility that produced the viral vector lot....Pennsylvania Vector Core is (as of April 2016) also performed by ddPCR.  Quality Control Addgene ensures high.... To learn which specific QC experiments were performed on your lot, please contact us. Full sequencing...the Viral Genome Next-generation sequencing is performed on viral genomes isolated from the final AAV preparation...

...containing full information about the suggested gRNAs to use, including all information provided in the...phenotypic readout, similar mapping could be rapidly performed by targeting all potential components of a signaling...available through Addgene. This means you could perform a screen with one library from Addgene, and then.... Thus, it is always a good idea to keep a bioinformatician close if you need help putting your list together... in vitro, but there is enormous potential in performing in vivo screens, studying genes affecting cellular...several algorithms. Please read the “Detailed Information” text linked to each reference library before...adapters. Output_UserInputParams: Contains all the information that you have input in the web interface, making...

...repository that can be used to perform screens in human cells. General information regarding how to use a CRISPR...assembly cloning platform called GMAP (Gibson assembly-based modular assembly platform). GMAP is based ...-wide Screening Using CRISPR/Cas9”. Detailed information regarding the Wu Lab library can be found in ...orphan GPCRs demonstrating the utility of this platform for drug discovery. Researchers can use plasmids...stop codon) and closed (including a stop codon) formats. The collection is compatible with both standard...Gateway cloning and the FNLCR Combinatorial Cloning Platform (CCP); the latter of which greatly enhances the.... PubMed PMID: 26165496 GMAP - A new assembly platform for generating both simple and complex DNA constructs...

...accepted as a grad student. I chose to focus on misinformation about sexual and reproductive health, as the...health issues creates conditions under which misinformation can flourish. I’ve never forgot about all the...perceive and evaluate peer-reviewed sources of information if those sources affirm or challenge their beliefs...term for a range of neat little tricks our minds perform to avoid cognitive dissonance (Kunda, 1990). More...conscious or unconsciously) pick apart sources of information they don’t like, looking for reasons to dismiss...scientific research is funded by the public and performed by researchers who received their educations from...schools and universities. However, much of the information gained from that research remains inaccessible...

...is, an antibody shown to perform well in one set of conditions may not perform as well in other conditions...assumed (Ayoubi et al., 2023). Additionally, their performance depends on the specific protocol used and the...For example, we have found that some antibodies perform well in a PFA-saponin fixation and permeabilization...population (Figure 1), further optimization can be performed to both maximize the separation and reduce any... is possible to design experiments where the performance of the antibody can be assessed through its ability...advantage of antibody validation efforts and information provided by external organizations like the Human...blood leukocytes. The file shared also contains information about the labeling pattern of several cell lines...

...the students performing these experiments have a good baseline for comparison. Performing WISH using these...are too few resources to teach students how to perform these experiments in the beginning. In most classrooms...observation and dissections. Students don’t usually perform more modern techniques used in genetics or molecular...addition, the Sánchez lab has made it easier to perform advanced developmental biology experiments in planarians...both WISH and RNAi experiments, as well as how to perform worm amputations. They also created an online companion...RNA interference (RNAi) is a powerful tool for performing targeted knockdowns of genes of interest and ...mixing with the liver paste, these E. coli are transformed with the appropriate plasmid and dsRNA expression...

...grade DNA Figure 1: Transformation introduces transformation grade plasmids into a bacterial cell...cell. Transformation grade DNA  Transformation is the process of inserting exogenous DNA in a microorganism...a plasmid preparation in 3 different ways: Transformation grade DNA Cloning grade DNA Transfection grade...plasmid DNA that is necessary for bacterial transformation of chemically and electrocompetent cells is...without commercially available kits and DNA for transformation doesn’t need to be as pure as for other uses...blog posts Learn about the differences between transformation, transduction, conjugation, and transfection...

... by polling and informal communications we made a decision to try out a Bioinformatics/HTS a journal club...popular, including those based on Twitter. At my former academic department, we had two recurring journal...teams had various levels of experience with bioinformatics and HTS, we all had much to learn. And, many...supplemental data, software repositories, or other information needed by someone to critically discuss the paper...Discussing next-generation sequencing at the Addgene bioinformatics journal club! What happens during journal...diving deep into the individual data points. All formats should be an open forum where people feel welcome...These questions should be tailored to the group and format but should almost universally include the questions...

...Molecular Cloning Technique Learn to Perform a Bacterial Transformation Deposit Your Positive Clones with...discusses some of the key things to consider when performing colony PCR.   Designing colony PCR primers ...downside: this type of primer does not provide information about the orientation of the insert. Orientation-specific...Orientation-specific primers: If you need information about insert orientation, then you might consider designing...each insert Backbone-Specific Provides information about the size of the insert and if it's in the...primers are plasmid dependent Doesn't provide information about the orientation of the insert Orientation-Specific...control. A good positive control is bacteria transformed with the same backbone plasmid. If this control...

... the pooled library is transformed into E. coli for amplification. Transformed E. coli are allowed to ...used to determine functional titer with a colony formation titering assay or similar protocol. After functional...library. This is similar to amplifying a plasmid: transform E. coli with the pooled library DNA, allow the...red.    Unlike a standard single plasmid transformation, though, a pooled library has hundreds to millions...diversity of the library sample, and any other information or scripts provided by the depositing lab for... be skipped or whether selecting colonies and performing Sanger sequencing will suffice…and the answer...mostlibrary requests specifically to allow scientists to perform functional titering and optimize transduction efficiency...

... line to perform downstream characterization of, isolating 2 or 3 distinct clones to perform key experiments...CRISPR-Cas system localizes to unintended sites and performs its programmed function there, this is an off-...associated with an elevated mutation burden. The CRISPR platform must be optimized to reduce off-targets and patient...Hopefully you can understand that depending on your platform, off-target effects should impact your experimental...would confound your experimental results and 2) perform targeted off-target measurements at those sites... to quantify off-target effects entirely is to perform whole genome sequencing. While this may be unnecessary... The con to these approaches is that NHEJ must perform the repair to allow for detection, not Homologous...

... understand that presenting your work in poster format differs in many ways from writing a manuscript....to include are a references section, contact information, and potentially a photograph of yourself. A ...where you can insert your references and contact information, and it is generally acceptable for the font ...accomplish this by employing a neat and organized format.  Columns. You can choose from either a two, three...than horizontally. Instead of displaying your information as columns, you may choose to display it as rows...barcode is included that links readers to more information if they desire it. Shapes. In this type of poster...poster, written and graphical information are inserted into larger circles or ovals, that are arranged...

... on advancing the paper-based platform toward a more field-ready format when the Zika virus outbreak was... 2014). The second was an in vitro paper-based platform that allowed gene circuits to be deployed out ...out of the laboratory in a cell-free and biosafe format that was stable for up to a year at room temperature...we say the timeline to develop our paper-based platform into clinically relevant diagnostics for the Zika...prototyping pipeline that can take pathogen sequence information from public databases to the production of validated...regions of the genome, we have deposited the best performing sensors and constructs encoding corresponding...and electronic companion technologies that has transformed the assay’s capabilities. Specifically this includes...

...are suitable for in vivo studies, our scientists perform all the standard quality control assays for AAVs... and Seqwell’s plexWell technology, we can now perform NGS at Addgene to sequence all the DNA packaged...should align to this reference sequence. Second, we perform a megaBLAST search on all the reads that did not...assembly of the unmapped reads, employing our bioinformatics software to assemble the reads into longer ...developed custom open-access Python scripts that can perform a deeper analysis of the data. Our serotype detection...contaminants in AAV preparations and provide information on these DNA species with exquisite detail (Lecomte...clinical-grade materials, and is not commonly performed for research-grade materials. Addgene is going...

...these types of experiments easier and faster to perform. Introduction to flow cytometry Flow cytometry ... path of a laser and the instrument collects information about the cell’s size, complexity and fluorescence...is captured by detectors and provides useful information about the cell. Light that continues in the same...is called forward scatter (FSC) and provides information about the relative size of the cell; larger cells...traveling is called side scatter (SSC) and provides information about the cell’s complexity. A cell with a high...antibodies to pass through the cell membrane. Formaldehyde fixation creates cross-links between lysine ...Sometimes, users will combine methods and fix with formaldehyde first to freeze everything in place followed...

... for accuracy and completeness. The information is then formatted and published as a report with a DOI...user-deposited data detailing how an antibody performed in an experiment.   Figure 1: The front page ...contains experimental data showing how the antibody performed in the submitted application, along with a brief... included in a publication, you'll find that information in the report as well.    Figure 3: An antibody... results of an immunocytochemistry experiment performed with the anti-Lhx6.1 antibody.    For a more detailed..., and a short description of how the antibody performed.  Figure 5: Results for the data report shown...

...activities (1). Researchers also discovered the modular format of some transcriptional activators: that the DNA...interactions. In later years, improvements to yeast transformation protocols also helped establish the yeast two-hybrid...  from Yu-Zhu Zhang’s lab) is an example of the former. In this system, histidine-auxotrophic host strains...applied to the bait in a single population (e.g. transforming both fusion plasmids, or mating two single haploid... haploid yeast fusion plasmid transformants into a diploid strain for screening), or arrayed on a 96 well... each well contains a defined prey fusion.  The former allows for screening greater numbers, but the latter...conducted and analysed with care to get reliable information. Although two-hybrid systems are a powerful tool...

...product is introduced into the cell by direct transformation. A given insert will typically contain both...selectable gene product for isolation of successful transformants. Addgene distributes several ready-to-use, modular...fluorescent tags in S. cerevisiae, looking at their performance in categories such as brightness, stability, ...associated with plasmid-based overexpression. Tim Formosa, at the University of Utah, has built a complete...from this collection will yield an insert with the format (protease site)-6xGly linker-(epitope tag)-ADH1...terminator-(selection marker). Additionally, Dr. Formosa has deposited six plasmids with a multiple cloning...modification in yeast developed by Dr. Pringle’s former lab at UNC Chapel Hill. Bähler et al.(2) describe...

...and drug development. Integrin structure and conformational complexity There are 24 known mammalian integrins...the cytoskeleton. Most integrins have three conformational states: bent-closed, extended-closed, and extended-open...further stabilizes the extended open integrin conformation. During embryonic development, extended-open...extracellular matrix. During immune responses, this same conformation encourages immune cell adhesion and migration...migration into inflamed tissues. Integrins' conformational flexibility is crucial for their function. With the...research reagents, researchers can measure this conformational change to investigate the role of specific ... Figure 1: Integrins can shape into three conformations: (A) bent-closed, (B) extended-closed, and (...

...Presented in figure 1). More information on Agrobacterium-mediated transformation can be found in this blog...modify several genomic loci at once (get more information on multiplexing gRNAs here). Cas9 is commonly...Simplified schematic of agrobacterium-mediated transformation in plant cells. Agrobacterium tumefaciens can...plant genome. B) The Ti plasmid and helper are transformed into agrobacterium tumefaciens and exposed to...methods do not involve double strand break (DSB) formation and do not require donor DNA, . These precise ...plant cytorhabdovirus as versatile expression platforms for planthopper and cereal genomic studies. New...Gelvin SB (2003) Agrobacterium-Mediated Plant Transformation: the Biology behind the “Gene-Jockeying” Tool...