...COVID-19 cases each week. Statistical Modeling Note -- If statistical modeling isn’t up your alley,...breaking the statistical independence assumption of the Poisson process. This modification also eases the...learning and statistical methods for fundamental scientific discovery and high-impact applications.       Alvin... of Chemistry and Chemical Biology at Harvard University; and Howard Hughes Medical Institute Investigator...months, pausing or terminating employment for a significant fraction of the workforce. Wet lab scientists...subfields.  Plasmid requests correlate with paper publication The dataset includes 11,426 plasmid requests ...14%). Spikes in plasmid requests correspond to publications of popular papers. Figure 1: Plasmid requests...

... be run in duplicate or triplicate. The closeness of the data points generated by replicate values provides... the standard dilution series in duplicate or triplicate. Replicates allows you to calculate the curve...blot. Standard curves are critical for a number of popular scientific applications, and the quality of a ... deviation of each replicate measurement divided by the mean of all the replicates multiplied by 100. ...regularly, and mix samples very well.  Use duplicates or triplicates to increase the accuracy of your experiment... value. Though it varies from assay to assay, typically the experimental value should be within 15% of...correlation between the actual values and the predicate values and measures how well the data points fit...

...caudal NST. (A-C) Low-magnification images showing viral spread. (D-F)  High-magnification images immuno-stained...vector performance. Yet, most of the technical data needed to replicate and build on others’ results is available...Produce Chemically and Functionally Distinct Vectors. Molecular Therapy - Methods and Clinical Development...and community concerns AAV vectors are complex biological reagents; many factors, like storage buffers ... right AAV for your system or to successfully replicate published works. Of particular importance to these...improve our Addgene Data Hub, 76% of respondents indicated they wanted to see more negative data (Figure ...out of fear it points to flaws or gaps in their technical ability. But, especially for AAV data, that may...

...and often cannot eradicate metastasis or eliminate the probability of a relapse. Chemical and radiation ...the tumor, this results in frequent physiological and morphological changes. Tumors differ in their genotypic...-level barricades put up by cancer. As mentioned in this article, cancers differ in practically every ...immune response to the development of tumors is a complicated and dynamic phenomenon that is interestingly ...either due to age, infections, or other strenuous biological changes. In contrast to the acute immune responses...themselves from attack. It is a region of incessant chemical activity, and consists of other cell types, especially...attack the cancer cells are also suppressed by chemical signals such as IL-8, IFNγ, and colony stimulating...

..., can be expressed from lentiCas9-Blast. Purpose of LentiCas9-blast LentiCas9-blast is a Cas9 expression...single-vector systems typically express Cas9 and a gRNA from the same plasmid. LentiCas9-blast virus can be...began with only a few inventory items offered domestically, but by the end of 2016, we expanded our viral...2016 were (drumroll please)... Top Lentivirus: lentiCas9-Blast Most of Addgene’s lentiviral inventory is... preps. Cas9 is the headliner of most CRISPR applications and can be expressed from your cell line of ...2014). It is a dual-vector system, meaning it is typically used in conjunction with a second vector that ...several benefits. First, the low viral titers typically associated with Cas9-expressing lentiviruses will...

...By Alyssa Neuhaus Genetic modification of microglial immune cells is critical for central nervous system...mice. B) Quantifications showing high microglia specificity of GFP expression for the indicated ssAAV5s...ssAAV5s. C) Quantifications showing high microglia transduction efficiency for the indicated ssAAV5s. Figure...Emily Bentley Building on their CROPseq method for optical pooled CRISPR screening, Paul Blainey’s lab has...compatible with enrichment, single-cell sequencing, and optical pooled screens in human cells. Addgene offers vectors...3’ long terminal repeats (LTR). The 3’ LTR is duplicated during lentiviral integration, producing a second...described in Anzalone et al., 2019, including modifications to the Cas9 or reverse transcriptase enzymes...

... University where she engineers genetically encoded voltage indicators. Tian developed a video-based high-throughput...red‑shifted archaerhodopsin‑derived genetically encoded voltage indicator which had improved performance in...the improved voltage indicator coming from the screen to understand mouse cortical and hippocampal neural... Haoyi Qiu He Tian Every two years, we seek applications for awards to early career scientists using ...virtual conference attendance. We also opened up applications to research technicians and undergraduate researchers...the brain. “We hope that iGluSnFR3 will enable significant steps forward in our understanding of synaptic...the chromophore that are susceptible to redox modification, the MPST hydrosulfide can be transferred to...

...in PubMed and choose the appropriate publication. This publication will be listed at the top of the final... "Published plasmids" or "Unpublished or pre-publication plasmids" and set a desired distribution status...quickly associate your plasmids with the original publication where you described them. Search for your article...plasmids. You can submit "Unpublished or pre-publication plasmids": Unpublished plasmids or plasmids that...accepted. Addgene also encourages submitting pre-publication plasmids so they can be available when the paper...preprint submission, we are happy to update the publication information once the manuscript is published ...your desired distribution status: Hold for publication: we will initiate our MTA process and perform...

...likely that the shifts caused to the practical and philosophical landscape of the scientific enterprise...Director of Media Communications, Harvard University Ray Howell -Founder of Howell Communications Philip Brenner.... Revitalizing biomedical research: recommendations from the Future of Research Chicago Symposium F1000Research...and keynote by former congressman and CEO of the American Association for the Advancement of Science, Rush...solutions. The event also culminated in a joint American Society for Biochemistry and Molecular Biology...union dues. Time will tell the extent of the implications of this ruling and whether graduate students...focus toward advocacy and action with sessions specifically chosen to better define productive routes for...

...Mechanisms of epigenetic modifications The main mechanism of epigenetic modification is chromatin remodeling...in this study,epigenetic changes were statistically significantly associated with reduced life-span, but...methylation in living cells." Journal of the American Chemical Society 126.19 (2004): 5982-5983. PubMed PMID...literally means ‘on top of the genome’). Epigenetic modifications can result in phenotypic changes without any...any change in genotype. Epigenetic modifications play a large role in turning genes ‘off’ and ‘on’, signaling...becomes. This prevents the cellular machinery from physically interacting with the genes in the compacted region...two main mechanisms, either through histone modifications, which can influence the transcription of genes...

...attractive choice for certain types of clinical or basic research applications. Considerations for IDLV use If...deficient lentiviral vector: prospects for safe clinical applications. PeerJ, 10, e13704. https://doi.org/10.7717...decades. Widely used across various research applications, they have become indispensable tools for manipulating... easily, without the need for time-consuming purification processes, and are powerful, due to their ability...into those of the cells they infect. These are typically DNA viruses, such as adeno-associated virus, herpesviruses...-lasting changes in cells. However, for some applications, integration might be considered a double-edged...plasmid contains Gag, Pol, Rev, and Tat genes. Specifically for our knowledge here, the Pol gene encodes...

...arrows indicate a conversion directly catalyzed by the base editor. Dotted red arrows indicate the change... point modifications. CRISPR base editing persists through multiple cell divisions, indicating that this...target modification in primary T cells, making them a promising tool for cell therapy applications. Dual...Base editing publication highlights In the table below, you'll find a list of publications we've highlighted...Early CRISPR applications were often limited by the low editing efficiency of homology-directed repair...all. Base editing to the rescue! Base editors chemically modify DNA bases without breaking the DNA backbone...following base editing and DNA repair. Black arrows indicate natural cellular processes. BER = base excision...

...which provides the AAV replication (rep) and capsid (cap) genes. AAV replication uses the host’s polymerase...this AAV purification video! It didn’t exist when I did my first iodixanol gradient purification, but I ...margins of my protocol. While these tips weren’t critical to the experiment, they definitely made my life...In this post, I’ll share some AAV production, purification, and titration tips, while also summarizing ...adenovirus genes which help mediate AAV viral replication; and 3) the transfer plasmid which encodes a ... that aids in the homologous recombination of duplicated regions like the ITRs. Restriction digest screening...consider performing PEG precipitation. Instead of sonicating the cell pellet, I lyse the cells with this AAV...

...blogger Jessica Polka, a Postdoctoral Research Fellow with Pamela Silver.  Most types of biological motion...these applications, purified R bodies could also perform as actuators in microelectromechanical systems... with new ways to interact mechanically with their environment.   Jessica Polka is a Postdoctoral Research...Systems Biology at Harvard Medical School. Follow her on Twitter @jessicapolka.     References 1. Kanabrocki...it to suit our purposes? In search of simple biological machines R bodies can repeatedly extend and...R-bodies, check out this video. Most types of biological motion (whether endocytosis, vesicle trafficking...to pH changes without the need for ATP or other chemical fuel sources. Like an actin filament or a microtubule...

...3’ end binds template DNA for PCR amplification, while modifications and homologous sequences are encoded...multiple modifications A key feature of IVA cloning is the ability to combine multiple modifications. Any ...’ Check your amplification by gel electrophoresis. In our experience, good amplification after PCR means...gain traction? It is likely that both vector amplification with low fidelity polymerases and the cost of...this approach requires no special reagents or purification kits, it is low cost and can be immediately ...to produce the linear fragments, as desired modifications and homologous regions can be encoded in the...parental DNA (37 °C for 15 mins). This enzyme specifically cuts methylated DNA, so it will selectively ...

... ensure they bind their antigens specifically. Possible applications for validation include western blot...addgene.org Addgene's Antibody Purification Protocol Addgene's Video Antibody Purification Protocol Addgene's Antibody...cells can perform human-like post-translational modifications that bacterial and most yeast cells cannot. ...production system may depend on what downstream applications you are planning, cost, and the amount of protein...secretes fewer of its own proteins, making the purification process simpler. Due to its high yields, E. ...using a mammalian production system, cell lines typically used include Human Embryonic Kidney (HEK) cells...production (Schwarz et. al., 2020). Harvest and purification At its core, harvesting recombinant antibodies...

...next-gen sequencing. ChIP is an incredibly useful application for exploring these interactions, and can give... protein interactions, including epigenetic modifications. But it's not an approach for the faint of heart...your sample before moving on to the next step. Typically, this can be done by cutting the tissue sample...into random, small pieces of roughly equal size, typically between 200-1000 bp. This is to ensure that your...background information. Your options for this step are sonication, which uses high-frequency sound waves to break...break up the DNA, or enzymatic digestion. Sonication provides a truly random digestion, but is harsher ...to the selective nature of the cutting. While sonication is usually the preferred method, enzymatic digestion...

...2024 American Chemical Society. Both PILATeS (18 kDa) and STReTCh (11 kDa) are significantly smaller...cofactors or using significantly truncated forms that limit its functionality. To make biophysical and structural...Sensors for Bioluminescent Readout of Mechanical Forces in Biological Systems. ACS Sensors, 9(7), 3489–3495...disruption in M. abscessus  Bioluminescent reporter of mechanical forces   Epitope tag antibody collection grows...performs well in many different antibody-based applications.  IPI’s recombinant version is a fusion of mouse...properties, and is amenable to a wide range of biophysical assays (Kroupova et al., 2024). They were able... Cereblon construct for crystallographic and biophysical studies of protein degraders. Nat Commun., 15...

...eukaryotic secretory pathway."Biochemical and biophysical research communications 430.3 (2013): 1114-1119. ...referring to the evil side of a mystical "Force," this quote is equally applicable to many microscopy experiments...within these compartments undergo significant post-translational modifications including glycosylation, disulfide... the cytoplasm? Or more troubling, what if a significant fraction of the fusion protein molecules fail...time. Previously, we have described a number of practical considerations that should be taken into account...human (and most eukaryotic) proteins localize to chemically distinct subcellular environments, including ...are equally susceptible to these non-native modifications, which can all potentially affect function [...

...fluorescent genetically encoded voltage indicator Article contributed by Shreya Vedantam Genetically encoded... for its biotechnological applications including conversion of CO2 into water and electrical energy generation...can be reconstituted in vivo to form a protein identical in sequence to the unmodified enzyme. Each half...tested the base editors in vivo and achieved therapeutically relevant editing efficiencies in the mouse ... base editors at Addgene. Levy et al., Nature Biomedical Engineering, 2020. PubMed PMID: 31937940. Detection...encoded voltage indicators (GEVIs) are proteins that detect and output the voltage changes in neural activity...fluorescent optics. Existing red fluorescent voltage indicators, however, fall short compared to GFP-based sensors...

...HIV. A notable landmark study showed significant clinical efficacy against a form of blindness known as...patients with clinical diagnosis of LCA and mutations in RPE65 (n=3), showed significant improvements ...and other ongoing clinical trials support the utility of AAV as a broadly applicable gene delivery tool...Almost all patients have achieved persistent immunological reconstitution with a normally functioning T...translating gene therapies from conceptual design to clinical trials involve identifying a therapeutic gene,...carcinogenic profiles have limited their usability in clinical settings. On the other hand, vectors derived from...Unlike most naturally occurring viruses used in clinical trials, wild type AAV is not associated with any...

...calcium indicator (Shemetov et al., 2021). In addition to FRET, intensiometric calcium indicators NIR-GECOs...characteristics, performance, and applications of near-infrared calcium indicators in this recent review (Shcherbakova...expertise is development and applications of fluorescent proteins, genetically encoded biosensors, and optogenetic...Qian, Y. et al. A genetically encoded near-infrared fluorescent calcium ion indicator. Nat Methods 16, ...Shemetov, A.A. et al. A near-infrared genetically encoded calcium indicator for in vivo imaging. Nat Biotechnol...Shcherbakova, D.M. Near-infrared and far-red genetically encoded indicators of neuronal activity. J Neurosci Methods...information on NIR FPs, their properties and applications. At the end, there is a Table that summarizes...

...covered in detail here. Classical double-strand break repair (DSBR) In the classical DSBR pathway, the 3’...not be immediately applicable in DSB-induced genome engineering, it has biological importance for the ... PubMed. A Mouse Geneticist's practical guide to CRISPR applications. Singh P, et al. Genetics 114.169771...pathways of DSBs, with a focus on repair for genome modification purposes. How does homology directed repair...and Homology Directed Repair (HDR). NHEJ is the canonical homology-independent pathway as it involves the... provided the DNA template used for repair is identical to the original DNA sequence at the DSB, or it...specific mutations into the damaged DNA if these modifications exist in the template DNA. There are several...

...holding a physical model of the protein in your hand – you will understand the power of a physical model.... researchers use physical models of protein structure? We have been creating physical models of proteins... your hand? At 3D Molecular Designs, we create physical models of proteins using 3D printing technology...and I encourage you to hold one. Why do we need physical models of protein structure? Structural biologists...biologists have always struggled to effectively communicate the nuances of protein structure in a flat 2-D...problem with the development of Byron’s Bender – a mechanical device that could be used to bend 1/8th inch ... example, when Tom Steitz was presented with a physical model of his large ribosomal subunit, he kept ...

...like team building, collaborative critical thinking, and communication among others. They additionally ...therapeutic and pre-clinical problems! Who knows, perhaps the ideas spurred by this “gamification” will lead to...to develop strategies to make our classes more practical and hands-on. Students in these courses get plenty...playing a card game called “Magic”. There were complicated rules that governed how this game was played....upon who your challenger was, you might have to drastically alter your strategy. I realized that similar ... as I try being compelling, interactive, and application-driven to boost curiosity, I have never experienced...engineering and strategic knowledge to solve a clinical disease case. As a teacher, you explain the game...

...presented chronologically. With a resume however, most often people choose a reversed chronological style ... job asks for experience in “statistical analysis”, mention “statistical analysis” (word by word) in your...together a grant application? Are you familiar with the publishing or patent application processes? How ... pages in a Curricula Vitae to showcase your publications, academic performances, and research experiences... applying for. Also remember that a resume is typically no more than 2 pages long. Write your resume for...information. Think "skills," not just "research and publications” What are the skills you developed through your...handled large data sets and are familiar with statistical analysis. If you have done fluorescent imaging...

...circle. Typically, a yellow check mark indicates that a particular parameter was automatically adjusted... The vertical black dotted line represents the cut site. Cutting and error-prone repair typically result...content. Many thanks to our guest blogger, Jessica Roginsky. Jessica Roginsky leads the scientific support ...This article was contributed by Jessica Roginsky, Scientific Support Lead at Synthego. Article source... of robust software tools. As a consequence, a critical step in the gene editing workflow - analyzing ...either one at a time or as a batch of hundreds, and indicate the guide RNA sequence you used. The ICE tool ...and interpret data about these types of edits is critical to the CRISPR workflow. For a multiplex sample...

...detection is crucial for clinical diagnostics, genotyping, and biotechnological advancements. Many methods...method can be used for an array of applications including clinical diagnostics (e.g. pathogen or virus...population for an array of molecular and clinical diagnostic applications. The company Mammoth Biosciences has...activated” Cas13 produces a quantifiable signal that indicates the presence of your targeted nucleic acid. To...amplified using RPA (recombinase polymerase amplification) or reverse transcriptase (RT)-RPA, respectively...RNA for subsequent detection by Cas13. This amplification step in combination with the ssRNA reporter ... introduced in 2017 so the lab made further modifications to improve their detection system. They named...

...preference.   Historically, ICSS was done by implanting an electrode and non-specifically electrically stimulating...the steps and technical considerations are directly applicable to the rat [6]. Technical differences include...A high-light sensitivity optical neural silencer: development and application to optogenetic control of...[2]. Laser activation is usually controlled automatically using motion capture software such as EthoVision...EthoVision. Consult the literature to find all the applications of optogenetics relevant to your field. Ultimately..., regardless of the type of behavior, certain technical considerations still apply. Testing stimulation...Inhibition of neuronal activation is a little more complicated since basal neuronal activity in your brain region...

...intuitive, or logical to a biologist that are not at all obvious, intuitive, or logical to a layperson...from other sources. Mode of communication Different modes of communication take up different amounts of...your audience needs to understand what you’re communicating. While I’m sure that there are loads of ways...new languages!) Jargon has a place in science communication, but the more you use, the higher cognitive ...This can be obvious, in that a lot of science communication is helping people understand new concepts. You...layperson. When you’re communicating ideas that you think are clearly connected, step back and ask yourself, ...pick and choose what concepts and details to communicate. It’s okay (and encouraged) to leave blank spaces...

...ability to specifically recognize and bind proteins. They’re used in a number of research and clinical applications... antibody’s ‘Y’. They are often used in clinical applications. Fab fragments can be created in lab in ...Shahied, 2004). They are often used for clinical applications, including imaging.  Diabodies are ~60 kDa...deeper penetration into a tissue, useful for applications like IHC, or be able to access an epitope folded...that would size-exclude an antibody. In imaging applications, smaller affinity reagents means higher resolution...Designed ankyrin repeat proteins, or DARPins, are genetically engineered small affinity proteins, around 14...proteins known as ankyrin repeat proteins. DARPins typically consist of 3-4 repeats, have high specificity ...

...conjugates (especially if you start to consider clinical applications). But there are a few other classes that... mechanisms. Most take advantage of chemical groups intrinsically present on antibodies, such as the primary...post-translationally. 4) Modifications made to antibodies, such as inclusion of non-canonical amino acids or peptide...Chemiluminescence, which is light produced by a chemical reaction, can be detected using specialized imaging...reactions produce a color change, so they are typically used in microscopy-based assays like immunohistochemistry...These properties allow for efficient signal amplification, making biotin conjugated antibodies good options...sensitive detection of low abundance targets with significant multiplexing potential. You will find antibody-oligonucleotide...

...Much of today's biological research requires a close examination of specific proteins within a system...system. This can be pretty complicated given that a single cell has tens of thousands of proteins functioning...single protein? Oftentimes, they rely on protein purification. In this strategy a single protein can be isolated...this blog will focus on one of the most common, purification based on the polyhistidine tag. What is the ...purity (Janknecht 1991, Hochuli 1988). In this purification approach, the metal ions are immobilized on ... incorporation of a cleavage site and a post-purification cleavage step to remove the tag in order to ...2000).  One downside of polyhistidine tag-based purification, however, is the propensity for nonspecific ...

...resistance cassette, and the R6K origin of replication to allow replication only in hosts carrying the bacteriophage...Belarusian State University Series of Physiological, Biochemical and Molecular Biology Sciences. 9.1 (...Lambda Red recombineering, which uses enzymes domesticated from the Lambda bacteriophage, is arguably the...enzymes can also accomplish the desired genomic modification. This technique is frequently called allelic...recipient of interest. Because pDS132 cannot be replicated in the recipient, the only antibiotic resistant...They hypothesized that genes that had evolved specifically for toxicity would allow stronger counter-selection...exchange. To do this, we performed the following modifications to pDS132 (Philippe et al., 2004): Replaced...

... brainstorming new technological innovations. Let’s take a look at three historical techniques: southern...computers, cell phones, and cars become more technologically advanced leaving earlier versions obsolete,...that save both time and money. Yet, knowledge of historical techniques comes in handy whether you are perusing...comparison to a control sample. Southern blotting applications range from confirming the results of cloning...cloning or knockout experiments to forensics and clinical diagnostics. One example of southern blotting being...interest by recording DNA abundance throughout the amplification process, rather than just at the end as in standard...fragments of a DNA sequence of interest to create a physical map of the DNA. Starting in the 1970s, restriction...

...of the FRET pair. Since many biological processes occur within the typical FRET range, FRET efficiency ...nanometers of each other, or measure the tension in the helical structure of spider silk, or the activity of a ...Fluorescence Resonance Energy Transfer. Unlike the typical excitation and emission of an excited fluorophore...donor fluorophore to the acceptor fluorophore. The typical steps in FRET are: 1. Donor fluorophore excitation...decision easier. 1. Müller, Sara M., et al. "Quantification of Förster resonance energy transfer by monitoring...Experiments using intermolecular FRET probes are typically used for studying protein-protein interactions...with an intermolecular probe is often either impractical (for biomolecules that are not proteins) or would...

...UCIBIO) focused on understanding the physiological and pathological roles of human cell glycans. Accordingly... biosynthetic pathway and have a wide range of clinical manifestations. PMM2-CDG is the most frequent ...reported that PMM2-CDG patients suffer from immunological defects. These are characterized by a predisposition...these lines to study how immune processes and biological pathways are altered in PMM2-CDG. These studies...reducing their risks for infection and other pathological conditions. References: Monticelli, M., Ferro... J., dos Reis Ferreira, V. & Videira, P. A. Immunological aspects of congenital disorders of glycosylation...Biology at the University of Colorado Anschutz Medical Campus in Aurora, Colorado, USA. The primary focus...

...supporters of science communication. To help you think about ways to effectively communicate your science, we...Scientist Jessica Welch Have Had to Explain Importance of Narrative in Science Communication Conversation...Conversation in Science Communication The Purpose of Science Communication #1 Mistake Scientists Make when Communicating...News at Harvard University. Effective science communication is required to promote public support for research... organization that works to promote science communication. Science in the News hosts a podcast, seminars...That’s right, #ActualLivingScientists, eager to communicate their science. In this first half of our two-...programs SITN runs and learn how they think about communicating science. Listen to the interview with Science...

...region you wish to stimulate 2) Fabrication and implantation of the optical fiber into the region of interest...run it? And, most importantly, what are the methodological and experimental limitations and considerations...numerous scientists at NIDA and some of the practicalities and considerations you may not find in the ...there of, depending on the opsin you use); the efficacy of optogenetics has already been proven. Therefore...region and cell type of interest is obviously of critical importance to any optogenetic experiment. There...pioneered the development of optogenetics, has genetically manipulated a number of the opsin genes to alter...optogenetics. Successful stereotaxic surgery relies on identification of the bregma and lambda sutures and subsequent...

...various time points. Measuring the optical density of the culture can indicate the number of cells present in...in-house protocol. DNA yields for this procedure are typically in excess of 100 ng/ul, more than enough DNA to...are, you’ll even have DNA left over for other applications, like PCR, cloning, transfection, or long-term...wash step or use a different strain. For most applications, Addgene uses and recommends DH5𝝰! This strain...cloning Bacterial strains adapted for protein purification use the cell’s resources to produce high levels...retransforming the plasmid into a strain meant for DNA purification. It sounds like extra work, but it will likely...your E. coli to ensure it is suitable for your application. We have previously reviewed many common lab ...