We narrowed to 1,042 results for: URE

...10hours. Temperature: FPs maturation times and fluorescent intensity can be affected by the temperature. For...no use to you. When using more than one FP, make sure their emission light does not overlap in wavelength...new GFP isolated from the Unagi eel was shown to mature independently of oxygen, making suitble for use...studies (24-30°C). Brightness: Brightness is a measure of how bright is the emission. Brightness is calculated...parameters (e.g. excitation light intensity, pH or temperature). Photostability: Fluorescent molecules gets ...lose the ability to emit light) after prolonged exposure to excitation light. Photostability can be as ...parameters (e.g. excitation light intensity, pH, or temperature). pH Stability: This parameter is important if...

...scientific community and their game changing nature. Full disclosure: Zimmer also spends some time on Addgene...Zimmer’s recent book, The State of Science: What the Future Holds and the Scientists Making It Happen, is an...I began my own career over 20 years ago, and I’m sure they were issues long before then. Other areas are...for me. A brief window in time “The most obvious future consequence of the undermining of the scientific...and technology.” This last section of the book, Future Science, describes the author’s thoughts on science... science and summarizing its past, present, and future in under 200 pages is an impressive task to set...Science could be a jumping off point to explore future rabbit holes. For the non-scientists in your life...

...inhibitors to your culture media prior to and after Cas9 introduction increases HDR, just make sure to remove ...distance. Homology arms and types of donors To make sure your donor molecule is used as the template, you...your Cas9 cuts in the coding region of a gene, make sure that the PAM edit you introduce is a silent mutation...exist as a repair template. To maximize HDR events, ensure cells are actively cycling through the cell cycle..., they will not undergo DNA replication, so make sure you are keeping an eye on those cells! To take this...mediates efficient homology-directed repair and termperature-controlled genome editing. Nat. Com., 8(2024...Resources on the Addgene Blog Check out our CRISPR Featured Topic Page Learn about Non-Homologous End Joining...

...different systems could be combined effectively (3). In Figure 1 for instance, the DBD of the fusion protein shown... interest(POI’s) are fused to TAs and DBDs (see Figure 2a) (4). These fusions are then co-expressed in...DBD-POI fusion (commonly referred to as the “bait”, Figure 2b) cannot activate reporter gene expression on...promoter of the reporter gene to activate expression (Figure 2c). The reporter gene then allows detection of...the HIS3 gene, which is required for histidine manufacture and therefore growth on his-deficient media. ...known to bind serves as a good positive control to ensure that your setup works. The last thing you want ... system to detect protein-protein interactions. Nature. 1989; 340:245-246. PubMed PMID: 2547163. 5. Ozawa...

... Addgene scientists inspect each sample and make sure they have been barcoded with a unique ID number ...number (1D barcode as shown below). This number ensures that every sample can be tracked easily at each step...involves good aseptic technique and controls to ensure that nothing is contaminated. We use filtered pipet...both of these are scanned into our LIMS system to ensure that they match, helping us to avoid mixing up ...glycerol stocks on Fridays. From each overnight culture, we prepare three sets of 2D barcoded tubes. Two...receiving, transforming, and storage process, we ensure that the sample tubes that we use match those of... to help track samples. They will help your lab ensure that sample ID’s are unique, consistently labeled...

...systemic biases which, in the absence of support and structure, prevent science from attracting and keeping diverse... diverse talent in science related careers. Failure to strengthen the talent pipeline sets the world up...to be successful after their training.   Figure 1: Professional Development Hub Workshop Attendees... with such a group dedicated to providing the structure and know-how to integrate career development in...areas of the scientific training and employment infrastructure. A Stakeholder Workshop at Janelia Research...2019 solidified the project and the findings were captured in this just released report “Enhancing Dissemination...attracting and launching science talent we must ensure that career skills training is a required and integral...

... everyone has their ideas heard.  So how can we assure that all participants feel comfortable sharing ...to encourage dissenting opinions is to foster a culture of inclusivity. Work with and value people who ...your lab/company/team is still working towards a culture of encouraging dissent, the next tip may be particularly...feel they can share opposing opinions as well. I am sure we have all experienced a meeting where one person...may not have the highest position or the longest tenure, but don’t let that stop you from speaking up.  ...them. If you can, set aside time to prepare for future challenges. This doesn’t have to be time-consuming...minutes every month dedicated to thinking about the future will improve your ability to innovate. (Finkelstein...

...Genome-scale measurement of off-target activity using Cas9 toxicity in high-throughput screens." Nature communications...this holds true in genome-wide screens.   Figure 1: Summary of the advantages of the new approach...effects of guide expression and dsDNA breaks. Figure 2: Non-targeting vs. safe-targeting guides. The...dsDNA breaks caused by CRISPR. See supplemental figure 6 for more details. Safe-targeting guides also ...of off-target cleavage by CRISPR-Cas nucleases." Nature biotechnology 33.2 (2015): 187. Pubmed PMID: 25513782...nuclease specificity using truncated guide RNAs." Nature biotechnology 32.3 (2014): 279. Pubmed PMID: 24463574... Other Resources Watch Dr. Michael Bassik’s lecture on “Multiplexing with CRISPR Screens”    ...

...mCherry is constitutively expressed. Cells are then cultured and FACS sorted. Image from Mateo Sanchez and ...-7 Optimizing fluorescent tools for labeling structures and compartments in mammalian cells By Angela...fluorescent proteins are often used to label  specific structures, compartments, or specific localization of biomolecules...provide misleading information on where exactly a structure or protein is located within a cell. The Goedhart...encoded fluorescent markers for labeling specific structures within cells. HeLa cells were transfected...and mScarlet-I for labeling a list of specific structures within cells. For example, to improve nuclear...microtubules. Find these fluorescent mammalian cell structure markers plasmids at Addgene! Chertkova, AO, et...

...included a glossary at the end of this post! Figure 1: Base transitions are edits between the purine...proof of concept for further development. Figure 2: Mechanisms of deaminase-based adenine transversion...for base editor development going forward. Figure 3: Mechanism of gGBE. Created with BioRender.com...•G base editing in mammalian cells and embryos. Nature Biotechnology, 42(4), 638–650. https://doi.org/...editor by engineered human uracil DNA glycosylase. Nature Communications, 15(1), 4897. https://doi.org/10.1038... editor with an N-methylpurine DNA glycosylase. Nature Biotechnology, 41(8), 1080–1084. https://doi.org...efficient T-to-G and C-to-G editing in mammalian cells. Nature Biotechnology, 42(10), 1538–1547. https://doi.org...

...tools and infrastructure in use at research universities not only today, but also in the future. Reproducibility...choosing an ELN for your research, be sure to consider how its features do and don’t support reproducibility...without access to them, the record of research captured in the ELN is incomplete. Thus, to support reproducibility...for discussion and review. It’s also possible to capture Slack conversations and import them into RSpace...and workflows that went into it. The ability to capture the conversations and associate them with a particular...ability to deposit data that has already been structured in folders or notebooks in the ELN, without having...

...the success of a drug’s clinical and commercial future, and yet are incredibly difficult to find solutions...The consultants who enjoy their jobs most, get pleasure from taking up such complex challenges, quickly...parts of the job? With fast turnover of projects, pressure to maintain consistently high levels of performance...consulting can sometimes be challenging. The fast paced nature of projects often means working long hours both...using lessons from previous cases to better staff future teams. As with any highly collaborative work place... scientific background, training in reviewing literature, and critical analyses skills were applicable... and equity research firms, biotech start-ups, venture capital firms, and more. Essentially any role in...

...GFPs The availability of the three dimensional structure of GFP, its brighter mutant S65T-GFP, and the ...bfloGFPc1. The plasmids used for determining the structures are now available from Addgene - bfloGFPa1 and...Mitochondrial health, transport and turnover are important features to be monitored to study mitochondrial dysfunction...dysfunction. Easy-to-use tools to monitor these features were lacking until Roberta Gottlieb and Zhen Yan labs...pTRE-Tight-MitoTimer) to show its usefulness in cell culture to report changes in mitochondrial turnover and...against common tumor markers to enhance cancer cell capture. Fierer et al., PNAS 2014 Apr 1;111(13):E1176...molecules and exceed a physiologically relevant exposure in intensity and/or duration. Both of these strategies...

...something doesn’t work, it’s not a failure. It’s an education.   Figure 4: Zenodo page showing the IBEX...reagents that work for our experiments!)     Figure 1: The IBEX Imaging Community front page. That...both their successes (recommended reagents) and failures (not recommended reagents) along with information... directly from the Zenodo dataset.     Figure 2: Screenshot showing reagent entries with associated...discussion with others in the IBEX community.   Figure 3: Screenshot showing recent topics in the IBEX...method for high-content imaging of diverse tissues. Nature Protocols, 17(2), Article 2. https://doi.org/10.1038...

...here and no viral progeny are produced (Figure 1).  Figure 1: Comparison of the life cycle of a lentiviral...recombination or non-homologous end joining (see Figure 2).  Figure 2: Schematic representation of integration...cell but cannot integrate it into the host genome (Figure 1). IDLVs were first discovered by researchers ...continuously expressed (Fu et al., 2013). Using IDLVs ensures transient expression without the risks associated...titering IDLVs, there is no integrated piece to measure! Instead, two commonly employed methods for titering...induced by CRISPR-Cas nucleases in human cells. Nature Biotechnology, 31(9), 822–826. https://doi.org/...

...Spinach2 reveals the dynamic nature of trinucleotide repeat–containing RNA." Nature methods 10.12 (2013): 1219...generated allows for qualitative and quantitative measurement of processes including RNA movement by tagging...many structural properties with DFHBI, but its structure allows for greater contact with its RNA aptamer...in the orange or red channel, depending on the structure of the TO derivative (Dolgosheina et al., 2014...imaging small non-coding RNAs in mammalian cells." Nature communications 9.1 (2018): 656. PubMed PMID: 29440634...transcription using a photostable RNA–fluorophore complex." Nature chemical biology 13.11 (2017): 1187. PubMed PMID...efficient RNA mimics of green fluorescent protein." Nature structural & molecular biology 21.8 (2014): 658...

...insert into your vector of choice as shown in the figure below. To start, you need to have DNA fragments...enzymes is that they can all work at the same temperature, so the entire reaction takes an hour or less...an overlap of 15-40 bp, with a primer melting temperature greater than 48℃. Both Snapgene and NEB have ...fragments have stable single stranded DNA secondary structure, such as a hairpin or a stem loop (as might be...overlapping region between fragments also better ensures correct assembly order of the fragments as compared...’ exonuclease instead of using the exonuclease feature of T4 DNA polymerase, which must be controlled ... these kits may come at a high price, their manufacturers tout great efficiency and low time commitments...

...constant, or Fc, domain. Sure, we all know that the Fc domain provides structure, determines isotypes, and...does! While the Fc domain’s functions, besides structure and isotype determination, don’t generally affect...sorted into three main effector functions.  Figure 1: Fc effector functions in an immune response ...antibody-dependent cellular phagocytosis (ADCP). Figure adapted from van Erp, et al., 2019.    Antibody-dependent...description with the Fc domain as only providing structure and isotype determination, remember that it can...Ig Constant Region Effects on Variable Region Structure and Function. Frontiers in Microbiology, 7. https...

...through the La Vida Scholars summer program. I was unsure whether or not I wanted to major in biochemistry..., who let me know in March that I successfully secured an internship at Addgene! I was eager to accept...took into consideration just how busy I was and ensured that my decision deadline would be extended. I ...reached out to Amy with exciting news. I managed to secure a virtual summer program, teaching me the basics...each intern to complete and present on.    Figure 1: Addgene's summer interns presenting their capstone...crunch and knew that if we were to restart the procedure we would not have enough time. During the following...from the electrophoresis, my manager and I were unsure if there were any successful clones because the...

...model and a source of sdAbs (Figure 2) (Xu, et al., 2021).      Figure 2: Genetically modified mice...found similar antibodies in llamas and alpacas (Figure 1). At the time, researchers were already on a ...nanobodies (Arbabi-Ghahroudi, 2017)!      Figure 1: Comparison of a heavy-chain only antibody and... for in vivo experiments, due to their simple structure    sdAbs in research These advantages have contributed...Single-Domain Antibodies: Historical Perspective and Future Outlook. Front Immunol 8:1589. https://doi.org/...mice and llamas neutralize SARS-CoV-2 variants. Nature 595:278–282. https://doi.org/10.1038/s41586-021...

... a couple of lectures in the undergraduate pharmacology course, like one or two lectures a year. And then.... And make sure to subscribe to the Career Channel of the blog so you don’t miss the future posts in this... the first time. I walk[ed them] through their lecture, giving them advice, [and helped] proctor exams...how to design test questions, and how to design lectures. I really built a course from the ground up.”  ...careers while in grad school And just for good measure, here’s a few other posts about developing transferable...

...techniques with higher efficiency and simpler procedures have been developed. Some of these include, TA...annealed and ligated in vivo. Detailed FastCloning procedure To start the FastCloning process, the insert and...and the resultant products of the reaction in the figure from Chaokun et al., 2011 to the right. After PCR...isolated from methylation competent E. coli for this procedure to work properly. Directly transform chemically...chemically competent cells with the digested mixture. There is no need for further purification before transformation...vector + insert). You should screen 3-4 colonies to ensure that you find the correct product. Once you find...have a better understanding of the step by step procedure of FastCloning, here are some tips for high efficiency...

...carte." Nature Reviews Genetics 12, no. 4 (2011): 231-42. PubMed PMID: 21339765. Methods figures were adapted...purify reprogramming proteins. Table 1: Key Features of Different Methods for Generating iPSCs. One...even after many passages. Ban et al developed a temperature sensitive Sendai virus which is removed by culturing...endogenous pluripotency genes (i.e. Oct4) are other measures of reprogramming. If a retrovirus was used, then...field of induced pluripotency: Recent progress and future challenges." Journal of Cellular Physiology 228...pluripotent mouse stem cells by the piggyBac transposon." Nature Methods 6, no. 5 (2009): 363-69. PubMed PMID: 19337237...minicircle vector for deriving human iPS cells." Nature Methods 7, no. 3 (2010): 197-99. PubMed PMID: 20139967...

...been developed around how they replicate (Figure 2). Figure 2: Baltimore replication classes. Image credit...molecular biology tools, a very useful trait. Virus structure While all viruses have a genome inside a protein... with an envelope are called enveloped viruses (Figure 1). Like bacteria, only a tiny fraction of viruses...viruses are pathogenic (Balloux & van Dorp, 2017). Figure 1: A non-enveloped virus (left) and an enveloped...packaged in separate plasmids and expressed via cells cultured with the vectors. This allows the viral vectors...of Viral Vectors 101 posts about each type, so be sure to check our blog, or our Viral Vectors 101 eBook...

...never expected to be in this here and now. Our closure was earlier than most companies. Since the start...around the world mobilized on COVID-19 research, cures, and diagnostics. Almost immediately we began to...receiving packages. So the entire team mobilized to figure out how to deliver when labs finally opened.  I...Addgene’s Technology Transfer Team also moved fast to ensure that the majority of deposited materials to study...the dramatic reduction in distribution, we had to figure out a way to maintain our services during the crisis...accelerating research. Yet this was still not enough to ensure we could keep up with demand and keep new projects...scientific sharing of materials for the foreseeable future. ...

...expression of XFPs is possible by simply introducing a mixture of XFP genes. Stochastic multicolor labeling is...trans-activator Brainbow) (Sakaguchi et al., 2018) (Figure 1). Once introduced, fluorescent proteins were ...AAVs (prepared with Addgene #104110 - #104112) (Figure 2). We’ve also created constructs with containing...Kohl et al., 2014) (Addgene #104106 - #104108) (Figure 3) that can be introduced using in utero electroporation...samples and visualize. For more details on this procedure, find the protocol here.   Tips and troubleshooting... al. "Improved tools for the Brainbow toolbox." Nature methods 10.6 (2013): 540. PubMed PMID: 23817127...of fluorescent proteins in the nervous system." Nature 450.7166 (2007): 56. PubMed PMID: 17972876. Kohl...

...probe for imaging nascent and mature HA-tagged proteins in vivo. Nature Communications, 10(1). PubMed ...fragments (scFvs) (Inbar et al., 1972) are fused to mature fluorophores, and can bind and light up epitope...., 2004). This scFv hybrid provides the probe structure and stability from the scaffold as well as specificity...it is translated quickly by the ribosome. This feature provides HA frankenbody the ability to bind nascent... wide range of protein dynamics in vivo. In the future, this stitching technique could be applied to other...applications: CDR grafting to stable frameworks and structure-based framework engineering. Methods, 34(2), 184...

...make sure to rinse off the surface of the filter membrane with the surrounding solution to ensure excess...concentration to avoid over-diluting, and make sure to remeasure your titer after diluting to confirm. Luke...drastically with lower titers. As you can see in figure 2, the expression from the 5x1011 titer was essentially... in a roughly 6-fold increase in expression [3, Figure 2]. Note how expression increases exponentially...additional best practices. Keep your cells happy - Make sure your cells are as healthy as can be to prepare them...

...education for future generations. All suggestions, tips and critiques to improve the educational nature of this...in these courses get plenty of theory in their lectures, but it can be difficult to determine how much...hands-on, it will be “brains-on”! I know that even lectures should be “brains-on”, but as much as I try being...have never experienced 100% student engagement. I figured, an engaging card game could help me get the engagement...students to use concepts learned from previous lectures, articles and books. A game where they would use...spurred by this “gamification” will lead to new cures for disease someday! Accessing and modifying BUILD...

...be available from their original sources. Figure 1: Biological repositories that distribute mouse... such example is cell lines. The American Type Culture Collection (ATCC) is a private, nonprofit repository... community. Addgene was awarded a Fast Grant to ensure that the scientific community will continue having...have already been requested from Addgene. Figure 2: Total COVID-19 plasmids requested from Addgene...making materials available rapidly to scientists (Nature Biotechnology 2020). The continued success of repositories... repositories, during the pandemic and in the future, depends on support from the scientific community...fundamental part of the scientific research infrastructure. References Chu H, Chan JF-W, Yuen TT-T, ...

...difficult if your work is based in the lab. If you’re unsure what to do during this period, check out these ... Organize data that you already collected into figures for papers or presentations. Learn a new skill ...novel findings. Most Micropublications contain one figure. If you aren’t there yet, write introductions to...while. Getting started early on introductions and literature reviews could make the months leading up to your...about reproducibility best practices Support your future experiments and other researchers by implementing...Also consider using this framework to design your future experiments.  Use Addgene’s AAV Data Hub. You can...can search the AAV Data Hub for tools to use in future experiments. If you’ve used Addgene’s AAV tools...

...Using Existing in silico Algorithm Infrastructure and a Vision for Future Enhancements. Frontiers in drug...immunogenicity score (Mattei et.al., 2022).    Figure 1: Generation of immunogenicity scores by predictive...individually-specific factors, such as previous exposures to that specific antigen, pre-existing conditions...factors that in silico prediction algorithms fail to capture. In vitro evaluation of ‌therapeutic immunogenicity...various components of the immune system. Figure 2: In-vitro models for immunogenicity prediction...and the advancement of precision medicine in the future.   Madhura Acharya is a former co-op and current...

...requires an NGG PAM to recognize the target. Osamu Nureki’s lab engineered SpCas9 (SpCas9-NG) with relaxed... with high precision - a valuable optogenetics feature when studying neuronal activity. The researchers...MerMAIDs plasmids at Addgene Opperman J et al., Nature Communications, 2019. PubMed PMID: 31346176. Using... of loops. These loops not only provide a 3-D structure for packing genomic DNA into nuclei, but are also...that heterodimerizes with CIBN upon blue light exposure (see Fig 1*). When the group targeted the enhancer...system promises to be a better model for the dynamic nature of genomic looping due to it’s briefer time scale...scale and its inducible and reversible nature. Find the LADL plasmids at Addgene Kim et al., Nat Methods...

...proteins and the potential for their work to enable future discoveries. Read on to learn more about Xin and...keep checking back in with the Addgene blog for future award opportunities. Xin Zhou: Fluorescent Light-inducible...active site. In these fusions, ultraviolet light exposure caused oligomerization of the Dronpa domains thereby...active site and shutting off the enzyme. Later exposure to cyan light stopped Dronpa oligomerization and...active site and still block enzyme function upon exposure to ultraviolet light. Previous iterations of FLiPs... needed an efficient and cost-effective way to measure the production of HMOs. Current HPLC-MS and fluorophore-based...ACS meeting. She hopes these biosensors and her future work with HMOs will enable researchers to engineer...

...from last meeting. Use some positive peer pressure to make sure everyone completes their assigned items ... on details that do not yet need to be decided. Figure out what you need to accomplish to get to the next...toward a solution. At the end of the meeting, make sure to thank everyone for their participation and offer... are working on. Building this into the agenda ensures that everyone gets a chance to talk and less vocal... so will the length of your group meetings. Make sure to keep them manageable. At Addgene, teams can add...all-company meetings, monthly still works best. Figure out what works for you for each of your meeting...

...synthetic) pFZ19 (yellow flame) Temperature inducible promoters Temperature sensitive expression systems ... expression at regular temperatures but can be induced by heat or cold exposure. Examples include the ...Types of inducible promoters Chemical agents, temperature, and light are all examples of factors that can...which a gene of choice is only expressed following exposure to a brief heat shock. In the case of Hsp70, the...since expression can be repressed by glucose. Temperature inducible promoters are also known for their ...

...surface of most immature B cells, its function is unknown. It has a monomer structure with a valency of... region: IgM, IgD, IgG, IgE, and IgA.    Figure 1: Diagram of an antibody labeled with Fc, Fab,...antigen of interest. Isotype controls are used to make sure the staining you see is due to target-specific antibody...region length. Now for some essential isotype nomenclature. Antibody heavy chain proteins as well as the...cells begin by expressing IgM. IgM has a pentamer structure, which means each IgM molecule has a binding capacity... for IgM, IgD, and IgG are deleted.     Figure 2: Isotype switching requires DNA recombination...

... base-pair editing CRISPR reporters in the near future.  SRIRACCHA (stable, but reversible, integrated...reporter into the genome of mammalian cells.  Figure 1: The SRIRACCHA reporter construct. Image from... for assessing CRISPR/Cas9 activity Guide RNA structure and the site in the genome you are trying to target...Thus it is essential to identify the best sgRNA to ensure success in your CRISPR experiments. But how? There... fluorescence and in-frame fluorescence.  Figure 2: The GEmCherry reporter. Image from Højland Knudsen...FACs sorting, confirming yet again that sgRNA structure and genomic recognition sites play a large role... for editing, regulating and targeting genomes. Nature Biotechnology 32:347–355 . https://doi.org/10.1038...

...PACE) method requires constant turbidity of the culture, so bacteria must be grown in a turbidostat. ...activity degrades the displaced strand.   Figure 1: EvolvR consists of an error-prone polymerase...by 120-fold over background. Due to its modular nature, the Dueber Lab created a few versions of EvolvR... more than one genomic site for mutagenesis, a feature which they used to generate bacteria resistant ...at a user-defined loci in bacteria. Its modular nature provides versatility in defining the mutational...diversification of all nucleotides in a tunable window." Nature 560.7717 (2018): 248. PubMed PMID: 30069054. Sadanand..., Saheli. "EvolvR-ing to targeted mutagenesis." Nature biotechnology 36.9 (2018): 819. Additional resources...

...have stable ssDNA secondary structure — for example, the stem-loop structure of transcriptional terminators...higher temperature at which Gibson assembly takes place may limit formation of secondary structures at the...with 5′ overhangs generated either by annealing a mixture of products or from incomplete PCR — although higher...effective with as little as 3 ng of DNA!) Figure 1: Schematic of SLIC. To start the SLIC cloning...successful, but at a lower efficiency (~20%). Figure 2: Schematic of multicomponent assembly with SLIC...

...sandwiched between 2 loxp sites and is excised by Cre (Figure 1). This results in one loxp site and one lox2272...Since loxp and lox2272 can not recombine, this ensures that the transgene is no longer able to flip back...the cause of this spurious expression?   Figure 1.  Cre-recombinase binds to either pair of lox...orientation preventing the transgene from reverting. Figure created with BioRender.com   Causes of recombination...to flank the transgenes (non-homologous pairs) (Figure 2.). They found that almost all off-target expression... to exercise caution when using them.    Figure 2. Schematic diagram showing the mechanism of spontaneous...reduced reversion and less off-target expression. Figure created with BioRender.com   Strategies for...

...University of Manitoba. Location, Location, Location! Failure to Plan (for Storage) Is Planning to Fail Patience...Patience Is Bitter but Its Fruit Is Sweet The Future? I knew little about adeno-associated Viral Vectors...sufficient to overcome the transport issues. 2. Failure to plan (for storage) Is planning to fail Freeze...not take much time at all. Much of the previous literature using AAVs for DREADD delivery indicates ~7-21...to use AAV constructs with self-complementing structure (scAAV) (7). As single stranded DNA viruses, AAVs...from ~4.8 kB [8] to ~2.3 kB [9]).     4. The Future What new types of research may be enabled by the...experiments, a group of researchers has identified this feature of AAVs as a new tool for neuroanatomical tracing...

...Glutamate sensing reporters Abhi Aggarwal is a postbaccalaureate research technician in Kaspar Podgorski’s ...expression and trafficking of these biosensors.   Figure 1: Schematic of iGluSnFR3 (left). GluSnFR3 in pMinDisplay... domain), in electroporated in rat hippocampal cultured neurons, imaged 14 days post-transfection, excited...happen inside the cell,” says Pedre Pérez. Figure 2: Micrographs of the biosensor expressed in the...from opioid use disorder,” says Hubbard. Figure 3: 20x confocal image of the periaqueductal gray...response to light touch in mechanical allodynia. Figure 4: While Qiu’s work on the PV neurons is in progress...calcium, and membrane voltage,” says Tian. Figure 5: Citrine-tagged voltage indicator expressed in...

... a native peptide bond at the split site. Figure 1: The base editor is split into two plasmids and...the split base editors at Addgene. Levy et al., Nature Biomedical Engineering, 2020. PubMed PMID: 31937940...in higher quenching of mScarlet emission. Figure 2: Ace-mScarlet is a fusion between the voltage-sensitive...synthetic biology tools for cyanobacteria. Figure 3: Assembly of plasmids using the Cyanogate Kit...reducing Cre toxicity in long-term experiments.  Figure 4: The C-terminus of Cre (CreC) and the N-terminus...(PCB), is required for PhyBΔ to be functional. Exposure to red light induces PhyBΔ and PIF6APB binding...This novel tool is practical in cell and organ culture, as well as cell lineage studies. Eisenhoffer and...

...engineering enzymes, each with their own unique features and quirks. Here, we will review the genome engineering.... Dozens of variants exist with unique cutting features and evolved PAM sites, making Cas9 an adaptable...PAM does not require any G’s or C’s. One curious feature of this Cas is that once it is engaged by its target...target, it will non-specifically cut ssDNA. This feature is shared by other Cas enzymes outlined below as... proteins should not be overlooked. The unique features and targeting abilities of the other Cas family...Janice, et. al. Programmed DNA destruction by miniature CRISPR-Cas14 enzymes. Science, 362(6416): 839-...with the single-protein CRISPR effector Cas7-11. Nature, 597, 720-725 (2021). 10.1038/s41586-021-03886-...

...the analysis did find 263,000 more osteoporotic fractures. The overall economic savings to the US healthcare... Unfortunately, it also meant that, by the very nature of the way the experiments were performed, they...and cell research. Using cells from both sexes ensures that the diagnostics and therapeutic interventions...recent years, and you can see a summary of this literature in our health and medicine checklist. A common...not analyzed) may harm humans or contribute to failures in drug development – costs far greater than those...doing the research correctly from the get go. The future of sex and gender in research There is much work...Blog Learn How Scientists Are Trying to Shape the Future of Research Learn How Notions of Scientific Impact...

...individual parts are reusable, saving time and money on future assemblies. If you are new to MoClo, give our MoClo...synthesis of these molecules. To make large amounts of pure product in an affordable way, researchers test many...your yeast experimental goals are more general in nature, or you aren’t planning on using K. phaffii, then...proteins within the mitochondrial matrix.     Figure 1: A chart with MoClo kit recommendations for different...2023). The Plant Parts II kit has a focus on infrastructure with shuttle vectors, several delivery systems...targets or multiple sgRNAs against the same target to ensure efficient cutting. Beyond sgRNAs, MoClo can also...convolute the results of these experiments. The modular nature of the Fragmid system allows for direct comparisons...

...Behavioral experimentation I’ll discuss some of the key features of each of these steps. Today we’ll discuss selection...relies on identification of the bregma and lambda sutures and subsequent determination of the proper coordinates... your brain region of interest (as shown in the figure below). Since most of the precision of optogenetics...circuits underlying brain disease in animal models. Nature reviews Neuroscience. 2012;13(4):251-66. doi: 10.1038...of neuroscience. 2011;34:389-412. doi: 10.1146/annurev-neuro-061010-113817. PMID: 21692661.  3. Bernstein...direct comparative analysis of microbial opsins. Nature methods. 2012;9(2):159-72. doi: 10.1038/nmeth.1808...mice for light-induced activation and silencing. Nature neuroscience. 2012;15(5):793-802. doi: 10.1038/...

...Met protein complementation. In the procedure diagrammed in Figure 3, students first transform the S. ...available (6). Methionine synthesis in S. cerevisiae (Figure 1) occurs through a well-characterized pathway ... the deletion). Find the Plasmids Here Plasmid features enable multiple student experiments We used TOPO...coding sequences for ten different MET/Met genes (Figure 1) from the two yeast species into the pYES2.1/... controlled by the GAL1 promoter. The cloning procedure adds both V5 and His6 epitope tags to the C-termini...expression by western blotting in other experiments (Figure 2).   Complementation reveals conservation of function...

...(ITR) present at both ends.   Figure 1: A) The structure of an adenovirus: the virion is ~ 100nm...genomes/mL) can be measured by: Optical density: particle concentration is measured by looking at absorbance... construct from step #1 and allowed to stay in culture for up to 20 additional days at which time the ...recombinant adenoviruses using the AdEasy system." Nature protocols 2.5 (2007): 1236. PubMed PMID: 17546019...production of high-capacity adenoviral vectors." Nature protocols 4.4 (2009): 547. PubMed PMID: 19373227...