We narrowed to 614 results for: cat.1

...biological questions. The two most common uses are: 1) to test the expression level in a specific system...brightness is compared to that of EGFP which is set as 1. Some proteins are very dim (e.g. TagRFP657, which...Photostability can be as short as 100ms (EBFP) or as long as 1 hour (mAmetrine1.2). However, for most FPs it is a... to be considered when choosing an FP: Unique categories of fluorescent proteins Photoactivatable / Photoconvertible... used are GECIs - genetically encoded calcium indicators (e.g. GCaMP). Others include: pHluorin & pHTomato.... al. (2010) “Fluorescent proteins and their applications in imaging living cells and tissues”  Physiol...

...optimized for expression in E. coli: acoustic gene 1 (arg1) and acoustic gene 2 (arg2). The key difference... plasmids are available from Addgene! References 1 Bourdeau, R.W., Lee-Gosselin, A., Lakshmanan, A., ...Knowing where bacteria are located within their host is often key to understanding their role in both...reporters are not as well suited for imaging organs located deep in the body, like the gastrointestinal tract...lab recently discovered that gas vesicles also scatter sound waves and therefore produce signals in ultrasound... experiment to demonstrate the most exciting application for ARG’s: in vivo imaging. Specifically, arg1...

...mGold2s and mGold2t depending on the application. Figure 1: Super-resolution imaging of actin ...fluorescent proteins for extended imaging. Nature Communications, 16(1), 3241. https://doi.org/10.1038/s41467-025... high brightness and fast maturation time (Figure 1). See the paper for more details on small differences...fused to mVenus, mGold2s, or mGold2t. Bottom: Quantification of photostability. Image reused from Lee & ...available for tracking cell lineage, different applications require careful selection or design to ensure...for gene therapy and ex vivo cell engineering applications and is especially well-suited for lung cells...tool for circuit tracing and other neuroscience applications (Figure 4). Check out our blog post on AAV serotypes...

...focuses on the replication machinery of Kaposi’s sarcoma-associated herpesvirus.       References 1. Snapp, ...experiments or even negatively impact cells (see Figure 1). When behaving properly, FPs enable researchers ... disulfide bonds in the secretory pathway (Figure 1) [4,6–8]. Additionally, FPs that encode N-glycosylation...compartments undergo significant post-translational modifications including glycosylation, disulfide bond formation...are equally susceptible to these non-native modifications, which can all potentially affect function [...N-glycans [9]. Both of these posttranslational modifications can disrupt FP folding rendering the protein... non-optimized parental proteins. Our recent publication illustrated the quantitative increase in fluorescent...

...(lenti) Centrifugation Titration time ~1 day (AAV); ~1 week (lenti) 2–10 days Initial titers Higher...will help you decide which is right for you. Table 1: Mammalian versus insect cells for viral production...so most labs opt to purchase adenoviruses from dedicated adenoviral production labs or companies. Production...something you may have to optimize for your specific application. With mammalian systems, it is standard to do...more cells, while AAV can be titrated following purification using qPCR or digital droplet PCR. Insect cells...to collect your viral supernatant. Baculovirus replicates inside insect cells, and its lifecycle ends in...titer the virus, or go straight into your viral application, like protein production. Titration of baculovirus...

... Posts from Addgene 1) Twitter – Get your voice out there Here’s one thing you can do today: Sign up for...science communication are as varied as the field itself. Among other things, science communication comprises...certain aspects of art and education. If you suspect that science communication might be for you, don’t ... science education Do you aim to reach young people? Try networking with your local education community... Training – Get a formal education Some people get into science communication by taking a media or journalism...project. Science communication opens doors Getting involved in science communication has been the most...career researcher looking to break into science communication? Everybody goes about this in their own way....

.... Follow David on Twitter @driglar.   References: 1. Alberts, Bruce, et al. "Rescuing US biomedical research...Riglar Advocating for Science Symposium and Workshop 2016 – Sept 16-17 MIT, Boston The Advocating for Science...futureofresearch.org/advocating-for-science-boston-2016/. Sign Up For the Advocating for Science Symposium...Director of Media Communications, Harvard University Ray Howell -Founder of Howell Communications Philip Brenner...union dues. Time will tell the extent of the implications of this ruling and whether graduate students...labor laws, rather than as trainees in a special category, as has been the ‘labor-of-love’ philosophy of...tight lab budgets and even simply being able to categorize all postdocs at institutions, will be challenging...

...GpNLuc and OgNLuc are available through Addgene (Cat.# 70185 and 70186, respectively). Antares Chu, J...construct for Antares is available through Addgene (Cat. # 74279). Enhanced Nano-Lanterns  Suzuki, K., et...furimazine by Promega’s Chemistry group is a good story (1). From a deep sea shrimp to a compact powerhouse of...cellular analysis products at Promega.     References 1. Hall, M.P., et al. (2012) Engineered luciferase reporter...et al (2). In this post, I’ll cover two great applications of the NLuc technology. The first couples NLuc...fluorescence. The paper reports a wide variety of applications of the Enhanced Nano-Lanterns including monitoring...2017) Genetically encoded bioluminescent voltage indicator for multi-purpose use in wide range of bioimaging...

...tethered to bring them into close proximity (Figure 1). Formation of the complete FP results in a fluorescent...detected by fluorescence microscopy. Figure 1: How BiFC works. A split FP (e.g., yellow fluorescent... protein interactions in living cells. Nat Protoc 1:1278–1286. https://doi.org/10.1038/nprot.2006.201 ...PIPs, resulting in a fluorescent readout.   Applications for bimolecular fluorescence complementation...resulting from the assayed PPI. This high-throughput application is much more straightforward to carry out because...protein construction, cellular expression, and quantification (Kerppola, 2006, 2013). These are described... supraphysiological expression levels. BiFC quantification This is where BiFC outshines FRET — BiFC experiments...

...environments such as live cells and tissues (Figure 1). These flexible strategies give you the best of both...majority of in-cell labeling strategies have two parts: (1) a genetically encoded “tag” expressed as a fusion...Follow him on twitter @rhodamine110.   References 1. Liu, Zhe, Luke D. Lavis, and Eric Betzig. "Imaging...Subsequent advances in fluorescent proteins have replicated many of the properties once exclusive to small-molecules...phosphopantetheinyltransferase) – These enzymes catalyze the covalent attachment of a fluorophore ligand...labels for fixed-cell imaging, not for live-cell applications. Thus, these fluorophores are often bulky and...the Addgene Blog Learn About Other Protein Tags Catch Up on Your Fluorescent Protein Background Which ...

... fusions that retain their individual activities (1). Researchers also discovered the modular format of...systems could be combined effectively (3). In Figure 1 for instance, the DBD of the fusion protein shown ...ubiquitin system has been devised (8). References 1. Casadaban M.J., Martinez-Arias A., Shapira S.K., ... can often be accounted for using appropriate replicates and controls (see Tips below), and by validating...two-hybrid screening Minimizing false positives: Run replicates of the experiment to reduce the likelihood of...expression and correct folding, post-translational modification, protein degradation, access to the nucleus ...co-expression of the enzyme responsible for the modification in the assay host strain can help. In order...

...a rate of up to 1 per 500-1000 bp. In comparison, the DNA replication error rate is 1 per 109 bp. Unlike...variety of biological applications. First, GMAP-compatible backbones for lentivirus LV 1-5 and retrovirus ...Transcriptional Output-TANGO (PRESTO-TANGO) to screen the NCC-1 library of approved drugs against the entire kit; ...generated 5 common 30bp overlap sequences (Sites #1-5). Each overlap site encodes a unique restriction...is multiplex automated genome engineering (MAGE) (1). MAGE uses recombineering (2) to simultaneously incorporate...of GFP (SEP) by a 9 amino-acid linker (see Figure 1). The key to the biosensor lies in pH: DsRed is relatively...inactivate SEP and leave only DsRed to fluoresce (1). With variants that can be targeted to specific cellular...

...This gives each guide a score ranging between 0 and 1. The cutting frequency determination (CFD score) is...converted to a weight (which ranges between 0 and 1) so that they can be used in the rank equation above...converted to a weight (which ranges between 0 and 1) (shown in the graph below) so that it can be used...HDR. Optimal mutation in-depth One the most sophisticated features of CrispyCrunch is how it mutates guide...

...microscopic resolution for studying colloidal gold (1).  The method was based on using a thin plane (sheet...Siedentopf, and R,. Zsigmondy.  (1902) Ann. Phys.  315, 1 – 39.
 A. H. Voie, D. H. Burns, F. A. Spelman. (1993...microscopy/en_de/products/imaging-systems/lightsheet-z-1.html ...of LSFM microscopes (3,4). After this initial application, Stelzer’s group described the single-plane or...brightness with increasing depth caused by light scattering and absorption.  Finally, the photomultipliers...

...the predecessor GEVIs (called ASAP 1 and 2), they found an indicator that is optimized for 2-photon imaging...several other constructs in plasmid form. Fig. 1: Summary of the approach and outcomes of the screen...what you'll find in this post: Improved voltage indicator: JEDI-2P Novel class of light-gated potassium...-Myc [9E10] now available! Improved voltage indicator: JEDI-2P  by: Brian O'Neill  The François St-Pierre...have successfully developed an improved voltage indicator (GEVI) called JEDI-2P. Using a custom, high-throughput...screen for improved genetically-encoded voltage indicators (GEVIs) based on the ASAP sensors. The resultant...Sustained deep-tissue voltage recording using a fast indicator evolved for two-photon microscopy. Cell, 18, 3408...

.... In literary terms, conflict can be defined as (1) the struggle between two opposing forces or (2) the...journey even when they know the ending.    Fig. 1: We all know that the superheroes win in the end. ...formal scientific communications, they are fantastic fodder for science communication. The trick is to ... is far from the only way to communication science. Science communicators also use the hero’s journey,...this small glimpse into the world of science communication and can use what you've learned to share your...Finding Conflict in Science  In all science communication, what you are doing is telling a (nonfiction...details often aren’t included in scientific communications, but they’re firmly part of the story to the...

... Figure 1: SPARC and SPARC2 use the GAL4-UAS system along with precisely truncated attP sites to... several common genetic tools in Drosophila (Fig. 1):  GAL4-UAS, a transcription factor binding site....of PhiC31. In the SPARC system, attP sites are located on either side of a stop cassette, upstream of ...amount of labeling, you can alter the attP site. Truncating the attP site upstream of the stop cassette results...population is directly correlated to the length of the truncated attP site. The three SPARC variants, based on ...intermediate (d), and sparse (e) labeling of the calcium indicator, GCaMP6f, in the Drosophila optic lobe. Image ...Image from Isaacman-Beck et al., 2019. Applications for SPARC SPARC technology is versatile and can be ...

...target DNA quickly and accurately.  Figure 1: Steps in PCR reaction   This process of denaturation...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...

...into E. coli is a fairly inefficient process– just 1 out of 10,000 cells on average! Without some means...down especially fast and plates should be used witin 1 month for optimal efficiency. Beware of satellite ... perspective – a plasmid-containing cell must replicate the plasmid in addition to its own chromosomal...fully synthetic compounds. Antibiotics can be categorized based on whether they directly kill bacteria ...bacteriostatic); however, the distinction between the two categories may be a bit of a gray area as some bacteriostatic...Blocks 50S ribosomal subunit; inhibits aminoacyl translocation Bacteriostatic 50-100 ug/mL in EtOH Polymyxin...Binds 50S ribosomal subunit; inhibits peptidyl translocation Bacteriostatic 5-25 ug/mL in EtOH *In prokaryotes...

...and take a look at each of its parts. Figure 1: Parts of an AAV transfer plasmid. Parts shown in ..., ubiquitous expression, while the human synapsin 1 promoter is neuronal specific (Haery et al., 2019)...contains the proteins needed for the virus to replicate,  and the transfer plasmid which contains the ... by the AAV. On a plasmid map, its sequence is located between the ITRs. It can be anything you like as...expression of your transgene to a specific species or location. Promoters can also drive different levels of ...sequences that can flank the transgene and are located between the ITRs. Transfer plasmids often have ... the expression of your transgene. The WPRE is located at the end of the AAV genome, just upstream of ...

... similar antibodies in llamas and alpacas (Figure 1). At the time, researchers were already on a quest...nanobodies (Arbabi-Ghahroudi, 2017)!      Figure 1: Comparison of a heavy-chain only antibody and single-domain...composed of camels, llamas, and alpacas (and their undomesticated counterparts) - not exactly species that jump...to sdAbs becoming valuable tools for diverse applications. For example, RANbodies can be used as an alternative... excitement around using sdAbs in diagnostic applications. For example, fluorescently and radioactively...heavy-chain only antibodies. (A) Overview of genetic modifications made to generate the nanomouse. (VHH = variable...inspired this versatile family of tools, the applications they have already enabled, and the discoveries...

...more tools for plasmid mapping and design (Figure 1). Benchling: While you might think of Benchling as...Davis from the University of Utah.   Figure 1: A plasmid map created in SnapGene of Blue Flame plasmid...tackling these particular tasks, but they tend to be scattered about the internet. To help you keep your virtual...: Find sequences for plasmids assembled from publications and commercially available sources, as well ... as expression patterns, post-translational modifications and polymorphisms Codon optimization tool: To...making it ready to copy and paste into other applications Primer design tools Many of the above-listed... well as primer design for different cloning applications. Primer3 and Primer3Plus: For general primer...

...from distantly related organisms. In 2006, less than 1% of the genes deposited at Addgene were synthetic....synthetic biology is expanding rapidly.  References 1. Kunjapur, Aditya M., Philipp Pfingstag, and Neil ...published their results in a paper in Nature Communications announcing a new bioinformatics tool that can...at its sequence. MIT collaboration for gene classification This inquiry started when Neil Thompson, a ...have generated robust tools that have practical applications in monitoring the accidental or intentional ...from farther away in the tree of life." Nature communications 9.1 (2018): 4425. PubMed PMID: 30356044. PubMed...the lab-of-origin of engineered DNA." Nature communications 9.1 (2018): 3135. PubMed PMID: 30087331. PubMed...

...removing 1-20 nucleotides from the cleavage site specified by the gDNA. Depending on your application, this...About NgAgo User's First Impressions Here References 1. Gao, Feng, Xiao Z. Shen, Feng Jiang, Yongqiang Wu...template is supplied. Find Plasmids from the NgAgo Publication at Addgene Cas9 and NgAgo head to head NgAgo ...differences in cleavage efficiency. If this finding is replicated in larger studies, it will represent a key advantage...PubMed PMID: 27136078. Find plasmids from this publication at Addgene. 2. Swarts, Daan C., et al.  “DNA-... PMCID: PMC4697943. Find plasmids from this publication at Addgene. 3. Swarts, Daan C., et al. “Argonaute...

...organoid model (Sun, et al. 2021).  Figure 1: Example schematic from the Organoid EasyTag system...without altering that of the internal control (Fig 1). Because this method relies on the fluorescence ratio...be used to generate organoid cultures with a modification of interest. While the authors optimized the...mechanisms related to morphogenesis, cell fate specification, and human disease.     Sun, et al. bioRxiv...protoplasts, the enzymatic biosensor retains its catalytic activity.  As SLs are also known to trigger seed...parasitization of cereal crops. As a result, agricultural applications of these novel biosensors could have a major...

...B. Gurumurthy, and Masato Ohtsuka. Nat Protoc. 13(1) (2018):195-215. PubMed PMID: 29266098 Creating knock-in..., with protocols using dsDNA templates achieving ~1-10% efficiency. Easi-CRISPR uses a long ssDNA template...the gRNA design process and rules for various applications is necessary going forward. The standardization... I think I may have finally grasped just how complicated DNA repair pathways are. I fully admit to being...Alt-NHEJ), has been harnessed for CRISPR knock-in applications. CRISPR 101: Targeting RNA with Cas13a (C2c2...for CRISPR Genome Editing Other resources and publications CRISPOR gRNA design tool Researchers at the ...

...principles of fluorescent dyes/ligands Figure 1: Example structures of fluorophores, fluorescent dyes... dopamine D2/D3 receptors. Scientific Reports, 10(1), 21842. https://doi.org/10.1038/s41598-020-78827-...for bacterial imaging . Microbiology Spectrum, 12(1), e0369023. https://doi.org/10.1128/spectrum.03690...Biochimica et Biophysica Acta - Biomembranes, 1838(1), 15–33. https://doi.org/10.1016/j.bbamem.2013.09.005...emission wavelengths, which are pivotal in their application in biotechnology (Grimm & Lavis, 2022). These... substrates/peptides and antibodies. Example applications of this include coupling to an antibody, whereby...whereby excitation of blue light will indicate where FITC is bound through green light emission. Key considerations...

...antisense RNAs inhibit plasmid replication.   Figure 1: The replication problem. Iterons are repeated...actually does, and neither plasmid is maintained (Figure 1).  The partitioning problem Plasmid incompatibility...Agrobacterium tumefaciens. Genes & Genetic Systems 77:1–9 . https://doi.org/10.1266/ggs.77.1 Pubmed PMID: ...Origin of Replication? See our Plasmids 101: Origin of Replication blog).  The replication problem Plasmids...directly inhibit replication by binding to the origin of replication and blocking replication machinery (del... of microbial plasmids: classification and identification based on replication and transfer systems and...plasmid that contains the origin of replication and the replication control machinery (Need a refresher...

...By the end of 2023, our catalog had grown to 879 viral catalog items (Figure 1). The service caught on...over 100,000 viral vector preps!   Figure 1: A) Cumulative AAV preps available in the Addgene ...Neuromethods, 195, 77–101. https://doi.org/10.1007/978-1-0716-2918-5_5 Haery, L., Deverman, B. E., Matho, K...repository. GCaMP, a calcium indicator (biosensor, by Addgene’s categorization) was first developed in 2001....   Figure 2: Numbers of AAV tools, by category, available from Addgene as of 2023. A large... calcium and voltage sensors, are the broadest category of viral vector tools, which may help explain ...number of requests for vectors in the “controls” category, which includes things like viral vectors that...

...with a single insertion because it will segregate 3:1 in the T2 generation. You also have no control over... transgenic plants in the process.     References 1. Emami, S., Yee, M. and Dinneny, J. R. (2013) ‘A robust...transformation’, Plant Methods. BioMed Central, 2(1), p. 19. doi: 10.1186/1746-4811-2-19. PubMed PMID:...Agrobacterium tDNA insertion likely precipitated domestication of sweet potatoes (Kyndt et al., 2015)! It sounds...

...level of expression needed for most applications.   Figure 1: With blue light, the VVD domains of... that dimerizes upon exposure to blue light (Fig. 1). Another blue light-inducible dimerization domain...in @nchembio Title: Bye bye L-arabinose drive. Dedicated to Edo, Armin and @KhammashLab. I hope you all...

...following three strategies might help!     Figure 1: Species on species staining can be a problem because...same host species as your secondary antibody since 1) your secondary should not recognize Fab fragments... “species on species”) immunochemistry (IHC) applications an issue? The culprits are endogenous IgGs in...Partridge TA (1998). A New Blocking Method for Application of Murine Monoclonal Antibody to Mouse Tissue...

...determined LY6A, also known as stem cell antigen-1 (SCA-1), is the receptor used by PHP.B to reach the CNS...William L. Stanford. "Concise review: stem cell antigen‐1: expression, function, and enigma." Stem cells 25.6...Mbp region of the mouse chromosome. Of the genes located in this stretch of DNA, Ly6a seemed the most likely...

...acceptable. Aims should be mechanistic, doable by 1-3 people in 1-5 years, and contribute to resolving the overarching...possible, do at least two full length practices (~1 h) and get feedback. It’s also helpful to practice...anticipated outcome, and payoff for each aim. Learn to communicate your story with only the most salient details...

...guide RNA (sgRNA), provided it meets two conditions:  1) The sequence is unique compared to the rest of the...bystander bases” if you choose this system.  Fig. 1: Cytidine deamination takes place on the free strand...temperature-controlled genome editing. Nat. Commun. 2017 81 8, 1–9 (2017).Walton, R. T., Christie, K. A., Whittaker... than ever to make precise, targeted genome modifications. Cas9 has been modified to enable researchers...instead.  Choosing a Cas9 for site-specific modifications Site-specific deletions and knockouts Some experiments...mutation at a specific site e.g., deleting the catalytic residue of a protein or knocking out a very small...flexibility as to where your sgRNA needs to be located and there may not be a NGG PAM site anywhere to...

...oligodeoxynucleotides)) are commonly used for smaller modifications (~1-50 base pairs). Small edits require as little...Easi-CRISPR was shown to increasing editing efficiency from 1-10% with dsDNA to 25-50% with ssODNs in mouse models...ribonucleoproteins. Quadros RM, et al. Genome Biol. 18(1); (2017). PubMed.  Additional Resources on the Addgene...pathways of DSBs, with a focus on repair for genome modification purposes. How does homology directed repair...specific mutations into the damaged DNA if these modifications exist in the template DNA. There are several... genome engineering world and made targeted modifications feasible and even easy. This targeted-break ...that can be harnessed by researchers for many applications. Your favorite Cas enzyme, a guide RNA to your...

...sgRNA1), 2; darker than control, 1; very dark). Brighter fluorescence indicates a more successful gRNA design...She loves to communicate the latest and greatest in scientific content.      References 1. Cho, Seung ...In vitro screening for the best gRNA Figure 1: Validation of double strand break (DSB) mediated ...plasmid using standard cloning techniques (The XX indicates where your target is inserted, disrupting the ...effective ones to increase your chances of gene modification in vivo. Find Plasmids for Testing Your gRNAs...et al., 2014). The plasmid is sturdier than the delicate RNA and therefore there has less chance of contamination... the validation assay should give an accurate indication of how many pups will contain a mutation and ...

...Full "Applying for a Faculty Position" eBook Here 1. Have a competitive CV You don't necessarily need ...that people would agree is important to understand. (1-2 sentences). For example: Distinguishing correctly... What will be the focus of your first grant? (1-2 sentences) Simply stated, what are your aims... do need multiple first/corresponding author publications in quality journals. It helps to come from the...a job. The currency of scientific success is publications and grants. 2. Write succinct and easy to understand...humbling. My conviction that I had a flair for communication was rapidly dispelled. She would ask me, "Is...succinct. Slide decks leads to the opposite and can suffocate an exciting idea in a swamp of unnecessary details...

...Check out Addgene's New Viral Service! References 1. Hay, R.J., Macy, M.L. & Chen, T.R. “Mycoplasma infection...contaminations.” Methods in Molecular Biology 946 (2013). 1-13. Pubmed PMID: 23179822. 5. Segura, M.M., Kamen,...Production, concentration and titration of pseudotyped HIV-1-based lentiviral vectors."Nature protocols 4.4 (2009...positively charged polycation, polybrene. Interestingly, one study found that alternative polycations such as DEAE-dextran... freshly collected lentivirus for downstream applications. While this ensures that the highest titer virus...lentivirus may want to consider testing a variety of cation and serum sources to ensure the best possible transduction...establishes the best conditions for specific applications, stick with them! Changes in such variables ...

...animal will repeatedly nose-poke at the active hole [1, 2].    Place preference/place aversion also models... can be explained primarily in two different ways 1) your hypothesis is wrong or 2) there is some problem...follow him on Twitter @derekpsimonphd.    References 1. Stuber, Garret D., et al. "Excitatory transmission...EthoVision. Consult the literature to find all the applications of optogenetics relevant to your field. Ultimately...Inhibition of neuronal activation is a little more complicated since basal neuronal activity in your brain region...technology is improving and may one day reach the sophistication of mouse transgenics. Everything that has been... and memory in opioid addiction and science communication/science writing. Derek is also a pianist and...

...and is recommended for use in western blots (Figure 1). If you prefer to produce your own antibody or want...are also available from Addgene!   Figure 1: Experimental validation for Anti-V5 [IPI-SV5-Pk1]...biophysical studies of protein degraders. Nat Commun., 15(1), 8885. https://doi.org/10.1038/s41467-024-52871-9...performs well in many different antibody-based applications.  IPI’s recombinant version is a fusion of mouse...with additional cofactors or using significantly truncated forms that limit its functionality. To make biophysical...cellular processes in C. elegans: mScarlet3 for applications needing intense brightness (such as low-expression...luciferase fragment (expressed separately) assemble and catalyze the luminescence reaction (Figure 5). The researchers...

...sequencing. DNA replication by PCR has error rates that range from roughly 1 per 500bp to roughly 1 per 10 million...not cut within your insert Are in the desired location in your recipient plasmid (usually in the Multiple...amplify and design primers that will bind to and replicate it. The following image shows the ends of the ...Forward Primer will use the sequence 5'-ATGTGGCATATCTCGAAGTAC-3' for the region that binds the ORF and...primer, making our Forward Primer 5'-GAATTCATGTGGCATATCTCGAAGTAC-3'. Many restriction enzymes do not cut...final Forward Primer sequence of 5'-TAAGCAGAATTCATGTGGCATATCTCGAAGTAC-3'. For the Reverse Primer, the design...need to use the reverse complement to get PCR amplification. We can start similarly, taking the final 18...

...your budget, you should take the following steps: 1. Define your timeline Start with your thesis in mind...these decisions in the following order of importance: 1. First make decisions based on experimental timelines... several factors aren’t taken into consideration. 1. Always focus on the science When negotiating finances.... After identifying all the major equipment, categorize all of your minor tools, like pipets, or mini-centrifuges...titles in the regions where the host institute is located. For example, salaries in Kansas versus salaries... most start-up P.I.’s. It doesn’t have to be complicated, or difficult to create this list. Start BEFORE...discounts, collaborations, access to internal award applications, etc. If this is the institute that can help...

.../nmeth.2641. Epub 2013 Sep 1. PubMed. Find plasmids from this publication at Addgene. Scalable and...sequences are inserted into the genome. Figure 1: Self-excising cassette schematic from Dickinson et...genomic modifications, is very robust in C. elegans, and the methods for HDR-mediated modification continue...After self-excision, the resulting loxP scar is located in this synthetic intron, and only the fluorescent...use and highly efficient for C. elegans genome modification. Figure 2: Schematic from Dickinson et...permission from the Genetics Society of America. Applications of this system Dickinson et al. have developed...system could be used to make many other genome modifications, including targeted mutations. SEC-based systems...

...those ideas and build from those.   Figure 1: Virtual lab meeting? No problem! You can still creatively... on it – so it is important to take breaks. After 1 hour (or another designated time) of meeting, someone...Collaboration and creativity are pivotal in writing publications/grants too. As described by several works (Wuchty...backgrounds.” This work outlines the need for clear communication, fair credit through authorship, and acceptance...remotely. The authors stress the fact that “miscommunication, a lack of leadership, and inappropriate tools...approaches can lead to frustration, delay of publication, or even the termination of a project.” Take ...setting. Check out the blog post “Fostering Communication in Remote Collaborations” to get more ideas....

...sequences were initially discovered in E. coli in 1987 (1), the concept that these clustered repeat sequences...neutralize CRISPR/Cas. Earlier this year, expression of 1 of 5 anti-CRISPR genes were found to inactivate the...distinct types and subtypes using polythetic classification based on evolutionary similarity (5). Type ...2013 (7, 8). Nine months later, more than 1500 publications describe work to improve the tool’s specificity...various species and describe an assortment of applications. Despite the ubiquity of CRISPR/Cas toolkit,... multiple plasmids used for various platform applications. For more information on CRISPR/Cas, check out...for your research. Do you have a cool idea or application that involves CRISPR/Cas technology? Let us know...

... from mutations in the gene, and whether any publications have mentioned the gene. The Bloomington Drosophila...

...processed since the declaration, only a small fraction (<1%) of authors have asked for advice about the scope...where readers may locate data supporting the research reported in PLOS publications. The policy was not... is founded on the principle that scientific publications must contain enough information (or links to...probable to some extent - but that is the price of publication. Who is to say that the initial data-generating...logically annotated and arranged, as in an actual publication and not require interaction with the data authors...likely, at least in part, due to our inadequate cataloguing of how results were obtained and this alone is... Nature Publishing Group recently launched a publication called Scientific Data, which aims to act as ...

...and recorded as a DNA strand is synthesized (Figure 1). Sanger sequencing requires four separate reactions...trace” file to determine the sequence. Figure 1: Overview of Sanger sequencing technology. Created...Technologies. Current Protocols in Molecular Biology, 122(1). https://doi.org/10.1002/cpmb.59  Wang, Y., Zhao,...differences due to in silico variations and lack of verification. To address this issue, the QC team developed...Nanopore sequencing technology, bioinformatics and applications. Nature Biotechnology, 39(11), 1348–1365. https...

...distinguish between your two targets.    Figure 1: In the isotype conversion process the variable regions...natural isotype to that of a different group (Fig. 1). This process, called isotope conversion, gives users...published, knockout validated, and work in your application. But…they are the same isotype and therefore ...are identical for antibodies within certain classification groups called isotypes.  Why does isotype matter...