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  1. Finding and Joining Your Dream Lab

    Type
    Blog Post
    ...does one stand out amongst a pool of successful applicants and get chosen to work in his or her lab of choice...dream lab While none of us will feel completely happy at work all of the time, the goal is to spend as...the difference. Choose a lab that brings on the happy dance at least once in awhile. Acknowledgments...

  2. Communicating Your Science With Help From ComSciCon

    Type
    Blog Post
    ...interview or whether your receive the next grant you apply for. After all, the only value your work will have... slide, think about whether the same principle applies to other slides in your deck.  If someone asks ...stay tuned to our website and social channels to apply for our future events. In addition to this annual...

  3. Cas13d: Small RNA-targeting CRISPR enzymes for transcriptome engineering

    Type
    Blog Post
    ... exon skipping. Previous CRISPR exon-skipping approaches used two guide RNAs to remove a given exon from...proof-of-concept for pre-clinical and clinical applications of dCasRx. The identification of Type VI Cas13d...sharing will continue to grow, and we at Addgene appreciate our depositors for making their tools available...

  4. Developing a cold tolerant E. coli using specialized chaperones

    Type
    Blog Post
    ...can firmly gain a handle on understanding and appreciating their biology. The importance of psychrophilic... the empty vector. And thus was born ELSA - our happy lil’ cold-tolerant E. coli! Additionally, we’re ...re also working towards widening the applicability of our strategy using the pMBP-LbCas12a and pC013-Twinstrep-SUMO-huLwCas13a...

  5. Antibodies 101: Isotypes

    Type
    Blog Post
    ... and remove a pathogen. Different isotypes also appear at different stages of an immune response. There... isotype of an antibody so you can select an appropriate isotype control. An isotype control has the same... Isotype controls are used in antibody-based applications including flow cytometry, immunohistochemistry...

  6. Viral Vectors 101: Viral Vector Elements

    Type
    Blog Post
    ...infectivity. Next, the virus must assemble and have appropriate factors displayed on the outside of the particle... production, they can be helpful for certain applications and aid in ease of use. Our lentiviral guide...elements discussed are compiled for reference below. Happy virus making!  References and Resources Kulanayake...

  7. Viral Vectors 101: Biosensors

    Type
    Blog Post
    ...ready-to-use. While biosensors span the gamut of applications within chemistry, biology, and physics, in this... this blog we will focus on those which can be applied in cell culture and animal models primarily. Fun...Neurotransmission and Neuromodulation: Engineering and Applications. Frontiers in Cellular Neuroscience. 2019 Oct...

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

    Type
    Blog Post
    ...guidance and structure. I quickly learned that my approach needed to adapt based on individual needs. For... and disengagement. By tailoring my leadership approach to each person, I was able to create a more productive...tension between two team members with differing approaches—one detail-focused and the other big-picture ...

  9. Plasmids 101: Sequence and Ligation Independent Cloning (SLIC)

    Type
    Blog Post
    ...usable product with two 5′ overhangs, so these approaches require higher concentrations of insert to succeed...intermediate won’t just have nicks: it may have gaps or overlapping regions of single-stranded DNA. But as long as...of multicomponent assembly, as fragments with overlapping sequence homology can be assembled in a unique...

  10. Opto-Nanobodies: Using Light to Manipulate Cell Signaling and Protein Purification

    Type
    Blog Post
    ... protein purification with OptoNBs The first application for OptoNBs takes us in vitro: light-controlled...require a protein to be tagged, which limits these approaches to use with recombinant proteins. Purifying an...OptoNBs to control cell signaling The second application for OptoNBs takes us into cells: light-based ...

  11. A Deep Dive into BioBuilder

    Type
    Blog Post
    ...labs, BioBuilder inspires students and teachers to apply engineering principles to biological systems. Today...teams around the world who combine engineering approaches and scientific know-how to design/build/test ...students include The BioBuilder High School Apprenticeship, preparing local (Massachusetts), economically...

  12. Back to Bacteria: CRISPR gRNA Multiplexing Using tRNAs

    Type
    Blog Post
    ...) How many loci can Cas9 modify? To test the application of this system to gRNA multiplexing, Xie et al...multiplexing further illustrates the potential of PTGs. Applications of PTGs The major advantage of the PTG system...multiple gRNAs may be used to target genes with overlapping activity. Multiple gRNAs can also facilitate ...

  13. 3 Challenges in Plant Synthetic Biology

    Type
    Blog Post
    ...associated with molecular plant parts, particularly for applied projects intended for commercialization. Many of...patented, and building our own novel pieces where applicable. We will also be working with open source, transformation...but there are several hazards to navigate when mapping out a new project. We’ve shared some of the concerns...

  14. Will You Be My Mentor? Finding and Asking for Mentoring Support

    Type
    Blog Post
    ... I will describe some of the many ways you can approach finding someone to give you advice and guidance...see someone with a great professional presence, approach them for advice on what makes them that way. Is...organization has a mentoring program, consider tapping into it and helping make it work by being an educated...

  15. RaPID Detection of RNA-protein Interactions

    Type
    Blog Post
    ...work. APEX2, a faster peroxidase-based labeling approach, is an alternative option but it’s toxic to tissues...subtracted from Ramanathan et al.’s RaPID datasets. Applications of RaPID RaPID sounds like a nifty technique...help answer? Ramanathan et al. outline three applications of RaPID: Discovery. To discover novel RNA-...

  16. dTAG - You're it!

    Type
    Blog Post
    ...proteins is an exciting strategy, it is difficult to apply this technique to oncoproteins that lack selective...that are not possible with traditional genetic approaches. In addition, the dTAG system is versatile and...update with vendor and catalog numbers. As we are happy to share aliquots of dTAG-13 in the meantime, please...

  17. Hot Plasmids - March 2020 - base editors, GEVI, MoClo, and optogenetics

    Type
    Blog Post
    ... changes in neural activity when paired with appropriate fluorescent optics. Existing red fluorescent ...organisms such as E. coli and yeast. Because this approach uses standardized parts, it allows for greater...increasingly been recognized for its biotechnological applications including conversion of CO2 into water and electrical...

  18. Giving gRNAs a Facelift - Synthetic and Beyond

    Type
    Blog Post
    ...were developed before CRISPR-Cas9, for use in applications like siRNA and antisense oligonucleotides. Some... after less than a minute of exposure to the appropriate light. The photoactivatable counterpart to this...protocols to get your CRISPR experiment moving. Happy editing! References and Resources References Hendel...

  20. Celebrating 15 Years of Scientific Sharing

    Type
    Blog Post
    ... work on our external website appearance and functionality are happening and this strengthens our image...support more new projects. Diversification is happening. The Addgene team will have some fun challenges...

  21. Viral Vectors 101: An Introduction to AAV

    Type
    Blog Post
    ...experiments. Recombinant AAVs (rAAVs) The primary application of AAV is delivery of genetic cargo to a host...rates for AAVs are generally high enough for this approach to work, although it’s less effective in vivo ... S., et al. “Various AAV Serotypes and Their Applications in Gene Therapy: An Overview.” Cells, 12(5),...

  22. The time and cost required to make a plasmid

    Type
    Blog Post
    ...screens and the more traditional “one at a time” approach. According to his math, it takes five hours of...method is free for you, and many labs are quite happy to share, labs that get high volumes of plasmid ...or changes. Fun fact: Addgene’s high-throughput approach means we virtually always use every single well...

  23. Antibodies 101: Conjugation

    Type
    Blog Post
    ...especially if you start to consider clinical applications). But there are a few other classes that we ...proximity ligation, immuno-PCR, and single-cell applications (Hegazy, et al., 2020; Niemeyer, et al., 2007...al., 2015). However, there are certainly other approaches that involve modifying the antibody sequence ...

  24. Plasmids 101: The protein expression toolbox

    Type
    Blog Post
    ...permanent control mechanism? Where is the control happening? Is the regulation of an exogenously introduced...times) into the genome. Make sure to choose an appropriate promoter to express your cDNA, depending on the...flavors of activators which can be used with this approach - read more about them on our CRISPRa blog! ...

  25. Gendered Innovations: Why Does Sex of the Cell Matter?

    Type
    Blog Post
    ...and Western Europe have taken three strategic approaches to gender equality over the past several decades...Australia worked with stem cells in mice. They appropriately used male and female mice (using both sexes ... the sex of the most commonly used cell lines appearing in American Journal of Physiology-Cell Physiology...

  26. Teaching an Old DOG New Tricks: Controlling Protein Activity with GFP

    Type
    Blog Post
    ... Tang and Cepko were eager to see if they could apply GFP regulation to other types of proteins. They ...systems are easily adaptable to neuroscience applications, including optogenetics, and they should make...Tang note that their scaffold system is likely applicable to many types of proteins. For proteins with ...

  29. Split-BioID: An Improved Method for Studying Protein-Protein Interactions

    Type
    Blog Post
    ...biomolecules. They can be part of multiple dynamic and overlapping complexes that have distinct functions. Many .... (2017). Split-BioID a conditional proteomics approach to monitor the composition of spatiotemporally...S., & Varnaitė, R. (2016). Meet the neighbors: Mapping local protein interactomes by proximity‐dependent...

  30. Adeno Associated Virus (AAV) for Cell and Gene Therapy

    Type
    Blog Post
    ..., non-enveloped virus that carries a small, approximately 4.7 kb single stranded genome. AAV belongs to... over a period of 1 to 3 years – the treatment appears to be both safe and effective. Challenges for AAV...trials support the utility of AAV as a broadly applicable gene delivery tool, aspects of AAV biology should...

  31. A Look Back at One Year of Plasmid Sharing for COVID-19 Research

    Type
    Blog Post
    ...more about it on the blog. COVID-19 research is happening fast and we’re glad to join these efforts and ... JM, DiVenere AM, Kuo H-C, Javanmardi K, Le KC, Wrapp D, Lee AG, Liu Y, Chou C-W, Byrne PO, Hjorth CK,...resources on the Addgene blog Learn more about mapping the SARS-CoV-2 - human protein-protein interaction...

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

    Type
    Blog Post
    ...tail clip, or toe clip) and using PCR with the appropriate primers to detect your targeting vector or gene...your parental mouse should be hemizygous at the appropriate allele. When crossed to a mouse that is homozygous...F1 heterozygotes to each other will generate approximately 1 in 16 mice that harbor homozygous alleles ...

  33. Open Resources and Plasmid Tools For Studying C. elegans

    Type
    Blog Post
    ...Scientists are currently doing this using two approaches: single-cell RNA-sequencing to identify neuronal... 2019. For more information about the aims and approach of CeNGEN, check out their publication.   3’UTRome...that can be used to simulate muscle tissue as it applies to C. elegans locomotion. To see the other resources...

  34. Antibodies 101: Stripping and Reprobing Western Blots

    Type
    Blog Post
    ...protein in your sample? Or what if something weird happened during your western and your results look… funky...to probe the blot again, almost like new. This approach preserves your sample and saves you from having...retaining residual antibodies. If you have already applied antibodies and need to dry the membrane for storage...

  35. Plasmids 101: Modular Cloning

    Type
    Blog Post
    ...multigene constructs, through multi-TU assembly,happens via three steps: Level 0: Cloning of basic parts...multi-purpose it can be. Let’s review a list of MoClo applications. Genetic circuits Most things in biology are...phenotype is best described with “it’s complicated.” Applying MoClo Interested in using MoClo? You don’t have...

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

    Type
    Blog Post
    ...of competitive TED talk – to win the audience’s applause with your performance. When your institute opens...guided tours to show people where the lab magic is happening. 3) Volunteer – Step into science education Do... Blog - Hone your writing skills Take the DIY approach and start a blog. Stellar talents like Ed Yong...

  37. Protocol - Bacterial Transformation

    Type
    Protocol
    ... thaw on ice (approximately 20-30 mins). Remove agar plates (containing the appropriate antibiotic ) from...spreading device Reagents LB agar plate (with appropriate antibiotic) LB or SOC media Competent cells DNA...Pro-Tip Transformation efficiencies will be approximately 10-fold lower for ligation of inserts to vectors...transformation onto a 10 cm LB agar plate containing the appropriate antibiotic. Pro-Tips We recommend that you plate... take up the DNA, an electro-magnetic field is applied to the cell/DNA mixture to induce membrane permeability... to have access to an electroporator and the appropriate cuvettes. Follow the manufacturer's instructions...

  38. Protocol - pLKO.1 – TRC Cloning Vector

    Type
    Protocol
    ...your convenience. See “warranty information” in appendix. Table of Contents A. pLKO.1-TRC Cloning Vector...References H.1 Published articles H.2 Web resources I. Appendix I.1 Sequence of pLKO.1 TRC-Cloning Vector I.2 ...GenHunter: #Q401 FuGENE® 6 Transfection Reagent Roche Applied Biosciences: #11814443001 OPTI-MEM® serum-free ...protamine sulfate, and puromycin are located in the “Appendix”. F.2. Determining the Optimal Puromycin Concentration...overnight. Day 2: b. The target cells should be approximately 80-90% confluent. c. Dilute puromycin in the...increments. d. Label plates from 1-10 and add appropriate puromycin-containing media to cells. Days 3+:...overnight. Day 2: b. Target cells should be approximately 70% confluent. Change to fresh culture media...

  39. Multiplexed Capture of Promoter-enhancer 3D Chromatin Structures Using CRISPR

    Type
    Blog Post
    ... original CAPTURE and CAPTURE 2.0 methods.   Applications of CAPTURE 2.0 CAPTURE 2.0’s increased rate ... of chromatin interaction networks  A second application for CAPTURE 2.0 is the construction of spatial...temporal changes in chromatin interactions A third application for CAPTURE 2.0 is detection of temporal changes...

  40. Hot Plasmids and Viral Preps - May 2021

    Type
    Blog Post
    ...The CRISPR corner New ready-to-use viral preps Mapping antibody escape in SARS-CoV-2 by mutational scanning... sequencing. From Starr et al Cell., 2020. (B) Mapping antibody-escape mutations with the yeast display...specific anti-SARS-CoV-2 antibodies (Fig 1B). This approach can augment viral surveillance by enabling rapid...

  41. Fluorescent Proteins 101: Visualizing Subcellular Structures & Organelles

    Type
    Blog Post
    ... map the internal organization of a cell. This mapping can be done by using antibodies targeted to the...meaningful way it is important to choose the appropriate quantification methods and tools. It is usually...Pages 2. Cooper GM. 2000. The Cell: A Molecular Approach. 2nd Edition. 3.  CureFFI.org. Cell Biology 04...

  42. Truncated gRNAs for Regulating Gene Expression

    Type
    Blog Post
    ...virtually any genetic locus when complexed with an appropriately designed small RNA, or guide RNA (gRNA). The...expression with truncated gRNAs An alternative approach is to modulate Cas9’s nuclease activity by modifying...engineering parameters via a library-on-library approach." Nature methods 12.9 (2015): 823-826. PubMed ...

  43. Degrading DNA with Cascade-Cas3

    Type
    Blog Post
    ...Cascade binding and subsequent DNA degradation appears to be conserved. This DNA degradation portion can... portion of DNA using only a single gRNA. Cas3 appears to have fewer off-target alterations, likely due...Mashimo, T. (2022). Genome editing technology and applications with the type I CRISPR system. Gene and Genome...

  44. PRIDICT: Predicting Efficiencies of Prime Editing Guide RNAs

    Type
    Blog Post
    ...machine learning (PRIDICT, PRIDICT2.0).   How to apply PRIDICT in prime editing experiments Prediction ...pegRNA efficiencies with pridict.it. This approach leverages the strengths of PRIDICT/PRIDICT2.0 ... computational biology. His expertise includes applying machine learning techniques to make sense of biology...

  45. Tips for a 1st time AAV user (by a Rookie AAV user)

    Type
    Blog Post
    ...Addgene AAV Production protocol, I take a lazier approach for harvesting my AAV. I skip the PEG precipitation...the PEG will separate into two phases. If this happens, the solution can be cooled and the two phases ...immediately before iodixanol purification. This approach gives me a bit of flexibility since I can store...

  46. 3D Printing Meets CRISPR Cas9

    Type
    Blog Post
    ...of proteins. But they are still flat. To truly appreciate the 3D structure of these proteins, it is necessary...molecular viewing software.   In reality, this rarely happens. But if you ever have an opportunity to read one...invariably say something like “Oh,…I didn’t quite appreciate that….” For example, when Tom Steitz was presented...

  47. Plasmids 101: Optimizing Plasmid Yields

    Type
    Blog Post
    ...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...your E. coli to ensure it is suitable for your application. We have previously reviewed many common lab ...

  48. PITChing MMEJ as an Alternative Route for Gene Editing

    Type
    Blog Post
    ...(2015). “A Scalable Genome-Editing Based Approach for Mapping Multiprotein Complexes in Human Cells.” ...lies in the diversity of nucleases and their applications. It will be interesting to see what new editing...

  49. Tips for Using BLAST to Verify Plasmids

    Type
    Blog Post
    ...reference sequence into the Subject Sequence box that appears. Aligning two nucleotide sequences is probably ...frameshift mutation present in the ORF is readily apparent when viewing blastx results. When using blastx...webservices/#pairwise_sequence_alignment_psa Happy BLASTing! Do you have any tips for using BLAST to...

  50. Plasmids 101: Secondary Nanobody Toolbox

    Type
    Blog Post
    ..., Alexa Fluor 568, or Alexa Fluor 647 before application to the sample. Staining with this multi-color...Mouse IgG2a TP1129 pTP1005 (3x cysteines) Mouse Kappa TP1170 pTP1174 (1x cysteine) pTP1172 (3x cysteines...labeling for super-resolution imaging, rapid epitope-mapping and native protein complex isolation. eLife. PubMed...
Showing: 501 - 550 of 964 results