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  1. Unique and persistent IDs for improved reproducibility: Addgene now supports RRIDs and compact identifiers for all plasmids

    Type
    Blog Post
    ... section now lists the multiple PIDs separated by semi-colons. For our example plasmid it will read “lenti-Cas9...require authors to use specific PID formats. For example, several journals like eLife, Nature, and Cell ...these PID formats are new to you, we’ll go through examples to highlight the difference between them using...identifier and RRID formats. Let’s go back to our example plasmid, lenti-Cas9-VQR-Blast. If you scroll down...Bauer (Addgene plasmid # 87155 RRID:Addgene_87155)” Please note that we only provide recommendations for how...

  2. Virtual Science Conference Coming Up? Three Addgenies Share Their Experience

    Type
    Blog Post
    ...between sessions Like many large conferences with multiple sessions happening at once, virtual conferences...hurdle of getting up and asking in front of many people is lower,” says Bachle. (For presenters, there’...other ways to make your session more engaging, for example, by using short animations and visuals, or asking...make it seem more like you’re connecting with the people,” Abitua says. Poster sessions The poster session...be a “place” for networking within the app. For example, the NeuroSymposium had virtual tables that you...you can join to connect with others. People are already discussing at tables and the event hasn't even...

  3. Photosensitizer Induced Cell Ablation with FAP-TAP MG-2I-dL5**

    Type
    Blog Post
    ... a photosensitizer composed of the protein complex complex and the MG-2I-dL5** fluorogen that can be used...thus forming the full FAP-TAP complex (MG-2I-dL5**).   A FAP-TAP complex will light up if excited with...-TAP MG-2I-dL5** MG-2I-dL5** is a new FAP-TAP complex where dL5** is the FAP. It’s job is to bind MG-...fluorogenic dye.  MG-2I is the TAP portion of the complex and is derived from the malachite green (MG) dye...

  4. Visualizing Translation at the Single Molecule Level

    Type
    Blog Post
    ... requires extensive sample processing to avoid disrupting ribosome-mRNA complexes. Recent work has adapted...time. Both are helping researchers explore the complexity of translational control in cellular physiology...resolution of this system are achieved by using multiple copies of the PP7 and MS2 coat protein binding...constant tracking. You can then directly monitor multiple rounds of translation from this single mRNA; you...cases, creating a reporter mRNA requires inserting multiple hairpin sequences into the coding sequence and...

  5. Addgene Depositors Get More Citations

    Type
    Blog Post
    ...Table 1 provides an example calculation of the weights. Table 1: Journal Weighting Example Article Number...However, determining this exactly is difficult. For example, it is very plausible that science published in...research our adjustment controls for them.   So, for example, in 2010, 10 percent of the papers with Addgene...factors that could still be controlled for. As an example, plasmid-related areas of science could be growing...www.neil-t.com/ .   References 1. Cong, Le, et al. "Multiplex genome engineering using CRISPR/Cas systems." ...

  6. Four Factors that Differentiate the Stem Cell Field

    Type
    Blog Post
    ...which "profound" seems like an understatement. Multiple speakers from top minds in the stem cell field...just 8 years after the OSKM paper – as a prime example of the fast movement in this field.  As an Addgene...the details. The speakers at this meeting shared plenty of details, but they also uniformly stepped back...uncle with Parkinson's, a half-brother who is a quadriplegic due to a spinal cord injury, and numerous family...expensive Dr. Yamanaka explained that it took 50 people nearly 2 years to create the first stem cell line...not a rate-limiting step in stem cell research. People told me they couldn't have done their work without...

  7. Plasmids 101: How to Verify Your Plasmid Using a Restriction Digest Analysis

    Type
    Blog Post
    ...particular reference sequence if necessary. The example plasmid on the right has a total size of 7.3kb,.... Avoid star activity. Some endonucleases (for example BamHI) are capable of cleaving sequences which ... The glycerol in the buffer will make sure your sample settles in the gel well and the dyes provide a ...you to interpret the bands that you get in your sample lanes. Choose your ladder based on the expected...approximate the DNA mass of comparably intense samples of similar size. Run the gel at 80-150V until you...is not just necessary but also an easy process. Please vist Addgene's resource for plasmid verification...

  8. Using AAV for Neuronal Tracing

    Type
    Blog Post
    ...connections between different brain regions. For example, communication between the hippocampus and the ...dendrites of proximal neuronal cells. In the specific example of episodic memory, a subset of hippocampal neurons...for review see Dietzschold et al., 2008). Herpes Simplex Virus (HSV) also migrates through neurons where...that express Cre in only specific cell types, for example only dopaminergic neurons, neuronal tracing has...unique hue (plasmids at Addgene). A large-scale example of neuronal tracing with AAV vectors is the Allen...part of the brain in a less invasive manner . For example, in neuromuscular diseases it would be desirable...Labeling in the Phrenic Motor System of Adult Rats: Intrapleural Injection versus Intradiaphragmatic Injection...

  9. From Lab Bench to Leadership: Transitioning into Management as a PhD

    Type
    Blog Post
    ...motivations, strengths, and learning styles. For example, during my time as a lab scientist, I worked with... management. Invest time in understanding key principles through books, workshops, and online courses ...foster a cohesive, productive environment. For example, in my current role managing a cross-functional...facilitated a team meeting to highlight their complementary strengths. By encouraging alignment on goals... while changing industries and positions. Some people discover they truly enjoy management and choose...successfully transitioned to industry after his PhD and a couple of postdocs focusing on molecular biology and metabolism...

  10. Viral Vectors 101: Gamma-Retroviral Packaging Systems

    Type
    Blog Post
    ... cell research has prompted the development of multiple virus packaging methods. Across these methods,...transfection step of the viral packaging cell line. For example, Phoenix, a second generation γ-retrovirus packaging... to enter the nucleus through the nuclear pore complex. This feature may be useful for the transduction... the components found within the backbone. For example: pLXSN has the wild type LTR, an MCS for cloning...sequence. This post was written and updated by multiple Addgenies. It was originally published in 2018...  Multiple Authors...

  11. Hot Plasmids December 2018 - C. elegans RNAi, TurboID, Detecting Viral Outbreaks, and Protein Binding Specificity Tools

    Type
    Blog Post
    ...ever like to write about a recent plasmid deposit please sign up here.  Listen to the hot plasmids episode...to physically set up in the lab, RNAi is often completely ineffective or results in low penetrant phenotypes...low-cost, cell-free detection assay that can be completed without special equipment. What’s more, detection...encodes the LacZ𝛼 protein. Once expressed, LacZ𝛼 complexes with LacZ𝜔 and forms an active LacZ tetramer....red-b-D-galactopyranoside on the assay paper turning it purple, allowing for easy visual detection of the presence...modifying the sequence of the toehold switch. For example, Green has also developed toehold switches to detect...

  12. Building and Validating Recombinant Antibodies: A Case Study

    Type
    Blog Post
    ...working to visualize multiprotein complexes.  Fig. 2. Multiplex immunolabeling with subclass-switched...easily be engineered. Like humans, mice express multiple subclasses of IgG antibody. Broad range secondary...subclass specific secondary antibodies to detect multiple targets with different mAbs in the same experiment...secondary antibodies and subclassed switched R-mAbs, multiple targets could now be visualized in a single experiment...recombinant monoclonal antibodies for enhanced multiplex immunolabeling of brain. Elife. 2019 Jan 22;8:...

  13. Sharing is Caring: But How to Distribute Open Hardware?

    Type
    Blog Post
    ...achievement, the complexity of the device increases, as does the price. With this increased complexity comes the...very difficult to adapt the device to one's own samples and limits the use of microscopy in open science...teleporter," since a part developed in Germany, for example, can also be printed quickly in Madagascar. Platforms...a move towards more open research. The EU, for example, is calling for the opening of sources within the...hardware.    Conclusion Addgene is a great example of how sharing of data and molecular tools can ...

  14. Plasmids 101: Screens vs. Selections

    Type
    Blog Post
    ... range from a defined pathway or reaction to a complex event such as cancer or even aging. For both a ...differences in these approaches can be found. As an example, say you are considering either a cellular screen...libraries targeting all known genes. This is a perfect example of an unbiased, reverse screen – all known genes...possessing a specific mutation or phenotype. For example, a drug screen may be performed on a cell line ...patients may respond to drug treatment. Another example is using a mouse model of a particular disease ...that do not affect the function of interest. An example where a selection would work is if you are trying...

  15. A Practical Approach to Choosing the B(right)est Fluorescent Protein

    Type
    Blog Post
    ...you want to buy, it is worthwhile to test-drive a couple of candidates. Before you buy a new microscope,...microscope, it is smart to look at (and through) a couple of models. Before you start a new project with fluorescent...fluorescent proteins, the best advice is to try a couple of promising variants to check how they perform under...that are related to the imaging strategy and the sample. Practical brightness A protein with a high intrinsic...brightness. Consequently, it makes sense to identify a couple of promising fluorescent proteins and to determine... future application. In figure 2 we provide an example of the comparison of the practical brightness of...

  16. Enabling Precision Functional Genomics with the Target Accelerator Plasmid Collection

    Type
    Blog Post
    ...cancer precision medicine seems so simple! Take a patient’s tumor sample, use cutting edge genomic technologies...would be extremely helpful as a complementary approach. Also, in principle, these experiments could find...cancer phenotypes. These assays were designed to complement the one assay per gene approach that has been...genes. These should be used to create ever more complete “look-up-tables” of mutant function that could...

  17. Protein Tagging with CRISPR/Cas9: A Conversation with Mendenhall and Myers

    Type
    Blog Post
    ...This second plasmid saves you a couple of weeks. CRISPR saves you a couple of months. It’s higher throughput...system work? Mendenhall: The system is really quite simple but it offers improvements in efficiency and throughput...right thing to do to put these out there. It’s a simple solution. It would be silly not to share it as ...Since the Addgene page went up last month, over 340 people have signed up to receive alerts when plasmids ...additional tips, tricks, or words of advice for people who may want to apply this tagging system to their...

  18. Antibodies 101: Conjugation

    Type
    Blog Post
    ...antibodies present throughout our bodies carry out plenty of roles just the way they are, the research antibodies... the visible spectrum, so they are great for multiplexing and can be used in a variety of assay types....detection of low abundance targets with significant multiplexing potential. You will find antibody-oligonucleotide...Fc-binding domain (Figure 1).  Figure 1: Examples of sites on antibodies used for conjugation. 1...from antibody to antibody or batch to batch. For example, take lysine conjugation, which is commonly used...reaction or the functionality of the conjugate. For example, Addgene’s antibodies are provided in a buffer ...

  19. Summer SciComm Series: Cognitive Load

    Type
    Blog Post
    ...listen quietly as people discuss their experiences. If you’re hearing multiple people with similar misconceptions... that a lot of science communication is helping people understand new concepts. You hopefully are aware...necessary” detail after “necessary” detail and end up completely overwhelming your audience - like the professor...group was just really tired and needed a slower, simpler presentation than I had planned; other times students...

  20. To Each HIS Own

    Type
    Blog Post
    ...   Fig. 1: A sample containing a polyhistidine tagged protein of interest (purple) and several contaminating...strategy a single protein can be isolated from a complex mix using affinity chromatography against an amino...metal ions are immobilized on a bead or resin and a sample containing the protein of interest flowed through...Ni2+-containing resin. The protein of interest (purple) binds to the Ni2+ and is immobilized along with...tags are also recommended if you need a very pure sample. Because binding strength is proportional to the...

  21. Choosing the B(right)est Fluorescent Protein: Photostability

    Type
    Blog Post
    ...fluorescent proteins can be directly compared. An example of a photostability measurement of various cyan...results in a loss of fluorescence, which, in its simplest form, can be described by a mono-exponential decay...intensity decay of a fluorophore does not follow a simple mono-exponential decay (Shaner et al, 2008; Bindels...applications in which far less power is used. For example you might find that you can get useful data from... diffusion, the proteins are (i) trapped in microdroplets in a water/oil emulsion, (ii) embedded in a ...fluorescent protein. In figure 1 we provide an example of such a comparison between cyan fluorescent proteins...

  22. Plasmids 101: The Wide World of Natural Plasmids

    Type
    Blog Post
    ...toxins are the clearest example of a plasmid benefiting its host. For example, cells with a penicillin...Shareck et al., 2004). Plasmid categorization is complex Plasmid classification has shifted over the years...Plasmids may also have more than one function: for example, Gateway cloning relies on genes found on an E....plasmids without clear analogues in nature. For example, helper plasmids were developed to replace helper... Multiple Authors...

  23. Mouse Modeling, Part 2: Breeding and Crossing Mice

    Type
    Blog Post
    ... parents. Here’s how to cross mice, using a simple example with the goal is to generate mice that have...genome, they are not commonly used to introduce multiple mutations. This is because as more mutations are...challenge that may arise when crossing mice with multiple mutations is if the mutations are on the same ...expressing strains Cre-lox strains are a common example of mice that are derived from crosses. In Part ...With this two-part guide to mouse modeling now complete, you should be prepared to plan an experimental...

  24. Science communication: 9 Strategies to Get Your Foot in the Door

    Type
    Blog Post
    ...public, for example at an anniversary or for Girls’ Day, offer guided tours to show people where the lab...into science education Do you aim to reach young people? Try networking with your local education community...Trust blog or the John Innes Student Voice are two examples. Many blogs like Plant Science today, Soapbox ...meetings. 8) Training – Get a formal education Some people get into science communication by taking a media...them to be the “be-all-and-end-all”. You can, for example, get an MSc Science Communication or MSc Science...

  25. Plasmids for Endogenous Gene Tagging in Human Cells

    Type
    Blog Post
    ... features of the gene intact. Here, we show an example gene targeted for tagging with GFP at the C terminus...interest. For a C-terminal tag, as shown in the example above, we inserted a linker and GFP tag at the ...the GFP tag precisely where we need it, in this example, at the C-terminus of the gene of interest. To ...CRISPR/Cas9 (Cas9 protein and the crRNA-tracrRNA complex) and the donor plasmid into the cells we use electroporation... Institute for Cell Science Plasmid Collection, please visit the Cell Catalog on the Allen Cell Explorer...hiPS cell lines generated using these plasmids, please visit the Cell Catalog on the Allen Cell Explorer...

  26. Tips for Technical Support Calls

    Type
    Blog Post
    ...buried beneath someone's lunch. Many times, I’m pleasantly surprised to discover that the most crucial information...from you and it’s wrong. Can you help us fix it? Please address as soon as possible!” and I always begin...your concern is based on data you have produced, please share the data. Maybe not all, but many bio companies...helping us understand the nature of your concern. Please be courteous. We understand how exasperating it...time and money on a potentially faulty product. Please remember that the person at the other end of that..., let your customer service agent know (nicely, please!). We can’t improve without feedback. And if you...

  27. Reaching out to China: Canton Nucleic Acids Forum (CNAF) 2015

    Type
    Blog Post
    ...Having never been to China myself, I was pleased to be able to participate when the organizers of the...Lowe lab have allowed them to begin understanding complex regulation of and by tRNAs and tRNA-derived RNAs...long hypothesized that life started with RNA and a simple cell membrane. His recent work shows that they ...replication with no proteins). The chemistry is complex and this work requires an interdisciplinary team... they do it. Of course, No conference would be complete without CRISPR Michael Terns from of the University...structural and functional component of the ribosome complex and Dr. Steitz quipped that RNA in the ribosome...

  28. Tips to Make the Most of a Scientific Conference

    Type
    Blog Post
    ...name for some context and maybe common ground. The people you meet may all be scientists but not all will...to practice beforehand. Generally, two types of people will visit your poster: those that want you to ...Try things like sitting at a table for lunch with people you don’t know or attending social events. I like...like to set goals for myself like meet ten new people each day. Exchange business cards. When you’re done...Follow up with those that you’ve met. Connect with people on LinkedIn or Twitter, for instance. If you’ve... head over to the “How to Make Friends and Meet People at a Scientific Conference” blog post. There’s ...

  29. Hot Plasmids and Viral Preps - May 2021

    Type
    Blog Post
    ...to co-localize multiple enzymes in a reaction pathway, the MPyV system provides a simple, efficient method...SARS-CoV-2 by mutational scanning By Seth Kasowitz As multiple antibody-based vaccines are deployed around the...many aaRSs exist within a multi-tRNA synthetase complex, the team investigated whether the generated antibodies...antibodies were able to immunoprecipitate the entire complex in three different cell lines. The Gräslund lab...jbc.RA120.01289     Novel protein cage provides simple enzyme protection in yeast bioproduction pathways...

  30. 10 Basic tips for mammalian cell culture

    Type
    Blog Post
    ... passage number. Especially in a lab where multiple people are working on different things, this will ...in use. Wipe the hood with a disinfectant (most people use 70% ethanol, but you can also use Incidin wipes... this purpose.Learn about the effects of CO2 on sample pH Cell morphology: Shape and size matter You should...which multiplies every 16 hours). So if you, for example, split cells on a Monday diluting them 1:10, you...contaminated.Learn how Addgene uses barcodes to track samples Keep track of passage numbers Keep an eye on the...

  31. Plasmids 101: Knockout/Knock-In Plasmids

    Type
    Blog Post
    ...through homologous recombination. The concept is simple: deliver a template with a modified version of ...homology is typical for targeting constructs. In the example shown in Figure 1, large sequences corresponding...is expressed (knocked in). As seen in the above example, you can remove a floxed resistance gene using ...plasmids described in the preceding sections are simple ways of knocking out a non-essential gene of interest...there are 3 loxP sites, recombination can occur in multiple ways. The desired recombination event will remove...tools for creating knockout animals or cell lines, please consider sharing them by depositing with Addgene...

  32. Genome-wide Screening Using CRISPR

    Type
    Blog Post
    ...strengthen your results. Enrichment or depletion of multiple gRNAs targeting the same gene can be strong evidence...jar of jellybeans, a pooled CRISPR library is a complex mixture. Pooled libraries are composed of many ...sorting and screened for a particular phenotype. For example, mutant cells can be used in drug screens to identify.... Loss of representation due to enrichment or depletion of specific gRNAs can lead to skewed results. ...any experiment, the use of appropriate controls, multiple replicates and several cell types can strengthen...

  33. Tips for Using BLAST to Verify Plasmids

    Type
    Blog Post
    ...getting-started guide for BLAST, which includes a simple explanation of the different BLAST programs, databases...consistently run our blastx searches with the “Low complexity regions” filter unchecked so that these regions...benefit is that blastp searches do not filter low complexity regions by default, meaning that you do not have...Clustal is our frequent choice for pairwise or multiple sequence alignments of nucleotide or protein sequences... sequences. We also use COBALT for aligning multiple protein sequences, particularly for comparing different...//www.ebi.ac.uk/Tools/webservices/#multiple_sequence_alignment_msa http://www.ebi.ac.uk/Tools/webservices...

  34. Which Fluorescent Protein Should I Use?

    Type
    Blog Post
    ...allowing very low background level of fluorescence. Examples for such photoactivatable or photoconvertible ... and their emission is green or red light. For example, T-Sapphire, LSSmOrange, and LSSmKate. Fluorescent...sensor), and iGluSnFr (glutamate sensor). More examples of these biosensors can be found at Addgene. ...also is also called Bimolecular Fluorescence Complementation (BiFC). 8 points to keep in mind when choosing...system can excite, and detect the emission. For example, if your microscope has only two lasers, at 488nm...minutes after it is translated to a few hours. For example, superfolder GFP (sfGFP) and mNeonGFP can fold ...

  35. Tips for arabidopsis transformation

    Type
    Blog Post
    ... of genes and proteins in living plants, to complementing mutant phenotypes. Arabidopsis is amenable to... sweet potatoes (Kyndt et al., 2015)! It sounds simple, and it pretty much is, but there are some tips...There are a bunch of great options, but here are a couple of my favorites. Plasmids from the Tsuyoshi Nakagawa...you transform early, your plants will still have plenty of time to produce seeds after you expose the flowers...for transformants. It’s very important to select multiple T1 plants from each transformation. I like to ... Agrobacterium T-DNAs with expressed genes: An example of a naturally transgenic food crop.’, Proceedings...

  36. Transferable Skills Guide: Identifying Your Transferable Skills

    Type
    Blog Post
    ...socks, persevered through a nerve wracking commute complete with a disabled train, and now you’re sitting ...within yourself, and to demonstrate them to the people who make hiring decisions. Though having all the...to other acquired skills.” It is nice when other people point out how awesome we are, but there is also...my wheelhouse”, take a second to reflect. For example, say your boss approaches your team with a project...front of a crowd and using your charisma to give people hope for some shiny vision on the horizon. You’...these responsibilities. If you are in charge of completing many meticulous protocols or tasks, you’ve probably...

  37. DNA Purification Without a Kit

    Type
    Blog Post
    ... takes 1-1.5 hours to complete, while this syringe-based method can be completed in 20 to 30 minutes. ...drawbacks of kits and typically follow the same principles: cells are lysed, RNAs and proteins are removed... centrifugation. At this point, the plasmid DNA sample can be treated with RNase to remove RNA contamination...waste footprint. The Zhao and Deng labs developed a simple method for regenerating silica columns that come... reduce the number of spin columns you need to complete your research. Do you know of other nucleic acid...

  38. Introducing Addgene's New Search - Find Plasmids More Easily For Your Research!

    Type
    Blog Post
    ...including depositing labs and institutions. For example, search for “Melina Fan” and you’ll be able to ...plasmids from a paper have been deposited? In this example we’re going to be looking for recombinant monoclonal...enough... If in the past you have tried to combine multiple terms in an Addgene plasmid search and found the...each filter is displayed in parentheses.  In this example I wanted to find a lentiviral plasmid to express...Starting with two search terms we see there are plenty of results, but I can’t browse through 300+ plasmids... plasmids that may be difficult to define for a simple search. These pages provide helpful information...

  39. CRISPR Between the Genes: How to Experiment with Enhancers and Epigenomics

    Type
    Blog Post
    ...arranged in a double helix. People are wrong. In reality, cellular DNA is a complex three-dimensional globule...Technique What It Locates ChIP-seq Proteins (for our example later, histones) ChiRP-seq RNA Hi-C DNA (genome-to-genome...appendix DNA is a 3D fractal). Enhancer biology is a complex and dynamic field. We’re going to focus on a tried... beyondthedish (left) and biobabel (right). People commonly think of DNA as a linear polymer of A,...Bedtools getFasta. These regions and their reverse complements were parsed for spCas9 PAM sites (NGG) and then...

  40. A Career in Grants: How To Become a Grant Professional

    Type
    Blog Post
    ...expertise on the processes needed to successfully implement, manage and close-out a grant. Grant professionals...reports are submitted and project evaluations are completed. To enter and advance the profession, you can ...  Skills such as research, collaboration across multiple disciplines, writing, project management, and ...application writing and development, project implementation, program evaluation and grant management.  ... time and resources. 3. Get a mentor, find your people Find someone who is working or has worked as a ...knowledge of the 8 core competencies through a multiple choice exam.  The bottom line on being a grants...

  41. Negotiating Work and Life: How to Find the Joy

    Type
    Blog Post
    ...to be managing. Being reminded of the seemingly simple actions we can take to help ourselves can be an...regular schedule at work. Choose a lab/job where people have regular schedules. Work 8 to 5 or 7 to 4 or...create “fake” family by forming a social circle of people in the same situation. Go to church, join a club... in the family to have to miss something – for example, say no to some of those birthday parties. Tell...print, but I actually let my daughter skip school a couple of times. Be sure to add time with your family,...time. For me, that’s been volunteer work at our temple, my kids’ school, and with professional organizations...

  42. New Tools Enable CRISPRa for Neuroscience Applications

    Type
    Blog Post
    ...Its RNA-guided nature makes CRISPRa much simpler to implement in genome-wide screens over cDNA libraries... Rochester Medical Center. During development, complex genetic programs specify and assemble diverse arrays...individual genes can be relatively laborious because multiple plasmids/viruses are needed to deliver all the... then packaged into AAV for in vivo delivery. Multiplex gene targeting is also possible and is only constrained...vivo simultaneous transcriptional activation of multiple genes in the brain using CRISPR–dCas9-activator...

  43. Viral Vectors 101: Pseudotyping

    Type
    Blog Post
    ...mechanisms behind the cytotoxicity can vary. For example, the popular VSV-G envelope protein causes cells...are differentially inactivated by nonspecific complement mechanisms in serum. Sometimes this is due to...cells that the viral vectors were produced in. For example, production of vesicular stomatitis virus, HIV-...sugars end up in the envelope and are targets for complement-based killing by anti-alphaGal antibodies. The...good at infecting most cells, but not all. For example, LDLR expression in hematopoietic stem cells, T... as described above and is inactivated by the complement system in serum (DePolo et al., 2000). Other ...

  44. Controlling for Off-target Effects with a New Genome-wide CRISPR Screen Design

    Type
    Blog Post
    ...al., guides were considered toxic if they were depleted from the screen. This toxicity was used a proxy... the growth screen, safe-targeting guides were depleted at greater rates than their non-targeting counterparts...Additionally, the distribution of enrichment and depletion scores for safe-targeting guides more closely ...noise, of dsDNA breaks caused by CRISPR. See supplemental figure 6 for more details. Safe-targeting guides...However, there was no significant difference in depletion of ricin sensitizer genes identified with short...Resources Watch Dr. Michael Bassik’s lecture on “Multiplexing with CRISPR Screens”    ...

  45. Avoiding the Dark Side of Fluorescent Protein Fusions with mOX FPs

    Type
    Blog Post
    ...combinations. For example, with standard fluorescent microscope filter sets, users can label multiple proteins ...often have absolute position requirements. For example, a KDEL ER retrieval sequence only functions at... even disrupt fusion protein localization. For example, the formation of inappropriate inter-chain disulfide...disulfide bonds between FPs blocks Golgi complex-localized fusions from properly exiting the ER and fluorescing... dark fluorescent proteins in your experiments, please consider the following: How can one ensure that...

  46. Changing Labor Laws Bring Increased Postdoc Wages

    Type
    Blog Post
    ...applying to, are doing - for example, your benefits may be changing. Please also feel free to contact us... salary was set at $47,476 and the date for implementation was set for Dec 1 2016. In order to be exempted...submitted letters to the Department of Labor (for example here and here) pushing for exemptions for postdocs...be on higher salaries (which have been made by multiple organizations across academia). Increased postdoc...wide consensus of academic stakeholders. There was ample opportunity to accept these recommendations, and...

  47. Tips for Using FRET in Your Experiments

    Type
    Blog Post
    ...revolutionised the way we apprehend molecular complexes and is still a very useful tool.    Like a guitar...technique, but it is very important to follow some simple rules to avoid common pitfalls and to use it at... These issues can be overcome by choosing a complementary pair of fluorophores. To maximize the FRET signal...especially when you investigate unknown molecular complexes. But it is a parameter that you have to keep in...most efficient approach for your own system. The simplest and the most popular one is the sensitised emission...photobleaching method The acceptor photobleaching method is simple but limited to a single measurement. This method...

  48. The Codon Guide to Synthetic Biology

    Type
    Blog Post
    ...foothills, I began to have conversations with many people who eagerly follow progress in synthetic biology...channels, books, podcasts, and communities to help people get started in synthetic biology. While not everything...NCI, NIH | License   I’m updating it regularly as people send me suggestions. The most useful parts of the...biology can be difficult to find. I hope that many people will find this guide useful as they start their...have additional resources that might be helpful, please reach out to me by email.  About the author Niko...

  49. Viral Vectors 101: Integrase-Deficient Lentiviral Vectors

    Type
    Blog Post
    ...across multiple plasmids. For simplicity, we’ll use the second-generation system as an example.  The ...Lentiviral vectors have been a staple in molecular biology for over three decades. Widely used across...double-stranded DNA transgene is incorporated into a large complex of both viral and cellular proteins and trafficked...the delivered payload is gradually reduced. For example, after using CRISPR/Cas9 to edit a gene, the expression...performing quantitative PCR titration of vector DNA in samples harvested 24 hours post-transduction (Wanisch and...

  50. Treating Muscular Dystrophy with CRISPR Gene Editing

    Type
    Blog Post
    ...hotspot for dystrophin, exons 45-55, there are multiple common deletions that maintain the protein’s reading...regularly. Dystrophin and genome editing Since complex oligonucleotide treatment comes with many challenges...skipping? For such a therapy to be successful, multiple requirements must be met. First, CRISPR would ...dystrophy. gRNAs can be designed to flank a single or multiple exons that, when deleted through NHEJ, will restore...therapy, it’s important to realize that DMD is a simpler case than other genetic diseases we’d like to treat...Thakore PI, Majoros WH, Reddy TE, Gersbach CA. Multiplex CRISPR/Cas9-based genome editing for correction...
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