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TypeBlog PostPublishedOct. 17, 2017, 12:55 p.m....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...
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Fluorescent Proteins 101: Visualizing Subcellular Structures & Organelles
TypeBlog PostPublishedJune 22, 2017, 2:30 p.m.... 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... -
PRIDICT: Predicting Efficiencies of Prime Editing Guide RNAs
TypeBlog PostPublishedJune 25, 2024, 1:15 p.m....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... -
Hot Plasmids and Viral Preps - May 2021
TypeBlog PostPublishedMay 18, 2021, 1:15 p.m....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... -
Truncated gRNAs for Regulating Gene Expression
TypeBlog PostPublishedJan. 10, 2017, 3:37 p.m....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 ... -
Plasmids 101: Optimizing Plasmid Yields
TypeBlog PostPublishedMay 26, 2016, 2:30 p.m....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 ... -
PITChing MMEJ as an Alternative Route for Gene Editing
TypeBlog PostPublishedFeb. 23, 2016, 3:30 p.m....(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... -
Tips for Using BLAST to Verify Plasmids
TypeBlog PostPublishedMay 29, 2014, 1:29 p.m....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... -
Tips for arabidopsis transformation
TypeBlog PostPublishedOct. 25, 2018, 1:23 p.m....Addgene Choosing an expression system Choosing an appropriate expression vector is the first step in generating...binary gateway vectors and can be used for promoter swapping and generating N and C terminally tagged fusion...phenotypes. For most phenotypic analysis it is appropriate to begin working in the T2 generation. The selection... -
Don’t FRET: Bimolecular Fluorescence Complementation Makes Visualizing Protein-Protein Interactions Easy
TypeBlog PostPublishedAug. 27, 2020, 1:15 p.m....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...without structural information, constructing appropriate controls may requires that you empirically test... -
Capturing a Moment with Marc Zimmer’s “The State of Science”
TypeBlog PostPublishedJuly 30, 2020, 1:15 p.m....What the Future Holds and the Scientists Making It Happen, is an exercise in restraint. The very first subheading... primarily on fluorescent proteins and their applications, tackles much broader issues in this brief volume...ago I would have nodded my head knowingly and appreciatively throughout this chapter. Now, these observations... -
AAVs CREATed for Gene Delivery to the CNS and PNS
TypeBlog PostPublishedSept. 28, 2017, 2:01 p.m.... an ideal way to deliver AAV to the PNS. It’s a happy accident that AAV.PHP.S targets the PNS since it...jumble of fluorescent neurons. A simple but flawed approach to multi-color labeling is to inject a mix of ...morphologic studies), as well as translational applications (mechanistic studies and gene therapy treatments... -
Summer SciComm Series: Modes of Communication
TypeBlog PostPublishedJuly 1, 2022, 1:15 p.m.... or burn yourself out trying to slog through unappealing work in your precious free time. And don’t be...- will be required to produce an outcome you’re happy with. Know if you’ll need specialized equipment ...then pick whichever is most accessible or most appealing to you. And if you’re not sure what all the possibilities... -
An Introduction to Adenovirus
TypeBlog PostPublishedOct. 3, 2023, 1:15 p.m....which a vaccine is available. However, here it appears that the culprit for these unusual cases was either... the removal of certain adenoviral genes that apparently contribute to virulence (Tollefson et al., 1996...on the Addgene blog Viral Vectors 101: Viral Applications Adenoviral Vector Production and Troubleshooting... -
Plasmids 101: Secondary Nanobody Toolbox
TypeBlog PostPublishedFeb. 27, 2018, 2:04 p.m...., 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... -
A Guide to Starting Your Own Journal Club
TypeBlog PostPublishedFeb. 6, 2020, 3 p.m....the most important is letting people know it’s happening. Aim for 3-4 weeks advance notice for the first...the Addgene bioinformatics journal club! What happens during journal club? There are many common ways.... These variations are suited for different applications and should be carefully chosen for each group... -
Plasmids 101: Colony PCR
TypeBlog PostPublishedMay 12, 2016, 2:30 p.m....primers. Find Empty Backbones at Addgene Each approach has advantages and drawbacks, which are summarized...which method you choose, make sure to use the appropriate antibiotic for selection. Lysing bacteria...to determine their size. Make sure to use an appropriate molecular weight standard for reference and to... -
CRISPR Antimicrobials
TypeBlog PostPublishedMay 3, 2016, 2:30 p.m....virulent or antibiotic resistant bacteria, and inappropriate antibiotic usage selects for resistant bacteria...to combat antibiotic resistance using multiple approaches. Major restaurant chains including Chipotle and...bacterial host range. To further potential clinical applications, Ando et al. recently engineered synthetic phages... -
Tips for Screening with Yeast Two Hybrid Systems
TypeBlog PostPublishedOct. 22, 2015, 2:30 p.m....or growth stage). A prey library can either be applied to the bait in a single population (e.g. transforming...although these can often be accounted for using appropriate replicates and controls (see Tips below), and.... Obrdlik, P., El-Bakkoury, M., Hamacher, T., Cappellaro, C., Vilarino, C., Fleischer, C., Ellerbrok, ... -
Cpf1: A New Tool for CRISPR Genome Editing
TypeBlog PostPublishedOct. 14, 2015, 2:30 p.m...., Zetsche et al. developed a Cpf1 multiplexing approach using a single crRNA array to express up to 4 ...-target editing in a cell culture model. The application of Cpf1 to genome editing is exciting both in...in terms of basic science and translational applications. This discovery of this type V CRISPR system ... -
The Importance of a Fun Workplace: Company Culture at Addgene
TypeBlog PostPublishedSept. 3, 2015, 2:30 p.m.... result seems to be a balance of productive and happy employees. The teambuilding culture helps us build...throughout the years. This team building merriment happened naturally when Addgene was a family affair back... sure people know that there is something fun happening! Addgene actually has chimes outside of our break... -
Deciphering the Mysteries of Behavior with Viral Vectors
TypeBlog PostPublishedSept. 17, 2019, 12:40 p.m....movement in an open field box or a circular corridor apparatus using a camera or tracking software. Locomotion...and might open new ways for a novel therapeutic approach against drug addiction. Anxiety tests Rodents...measuring the time spent in a particular part of the apparatus, it turned out that the IL-1 beta overexpressing... -
Rett Syndrome: A History of Research and Therapeutic Outlooks
TypeBlog PostPublishedNov. 19, 2020, 2:15 p.m....highlights some of the latest breakthrough therapeutic approaches. What is Rett Syndrome? Caused by a mutation ...technical challenges to overcome for both of these approaches, such as delivery of the editing machinery to...Extends the Survival of MeCP2-Null Mice without Apparent Toxicity after Intracisternal Delivery. Molecular... -
Tips and Tricks for Using Golden Gate Modular Cloning (MoClo)
TypeBlog PostPublishedJuly 8, 2025, 1:15 p.m.... process, and request your sequence with the appropriate flanking restriction sites and overhangs already... help you in your MoClo quest, so good luck and happy cloning! Resources Additional resources on the...Modular Cloning Plasmids 101: Modular Cloning Applications and Kits Plasmids 101: Golden Gate Cloning ... -
Plasmids 101: Shuttle Vectors
TypeBlog PostPublishedMay 20, 2025, 1:15 p.m....which helps increase cellular uptake, but this approach requires compatible cell lines that express viral...individual analysis. A shuttle vector designed for this approach in a mammalian cell experiment should still include...terms are typically used in different (though overlapping) ways. Broad host range plasmids are those that... -
Advanced Uses of Cre-lox and Flp-FRT - A Neuroscientist’s View
TypeBlog PostPublishedOct. 19, 2017, 1:54 p.m.... its reintroduction, inversion and re-inversion happen at the same probability. Continuous inversion of...the same time from two different plasmids this approach becomes problematic as it reduces the probability... Cell Types for Imaging and Long-Range Circuit Mapping.” Journal of Neuroscience 28(28): 7025–30. PubMed... -
Hot Plasmids Spring 2024
TypeBlog PostPublishedMay 23, 2024, 1:15 p.m....Paul Blainey’s lab has developed a multiplexing approach to Cas9-based pooled CRISPR screens called CROPseq-multi...for microglia research and broader therapeutic applications. Find microglia-targeting AAV plasmids here...reconstituting coding sequences for a range of applications, including CRISPR activation by dCas9-VPR, prime... -
AAV Titers: Where Do They Come From And What Do They Mean?
TypeBlog PostPublishedNov. 15, 2017, 3 p.m....used to generate AAV. One study showed that approximately 50% of viral capsids were genome-containing ...particles can be identified after negative staining and appear darker than full vector particles. This image shows...experiments? Because (physical) AAV titers are approximations and they depend highly on the experimental ... -
New Acoustic Reporter Genes: Ultrasound Imaging of Gene Expression
TypeBlog PostPublishedJan. 24, 2023, 2:15 p.m.... the bacteria were injected, limiting their applicability. They also did not scatter ultrasound nonlinearly...chromatic-opening agents to boost expression to appreciable levels. We wanted to create second-generation...ultrasound imaging of nanoscale acoustic biomolecules. Appl. Phys. Lett. 110, 073704 (2017). Hurt, R. C. et ... -
Changing Labor Laws Bring Increased Postdoc Wages
TypeBlog PostPublishedOct. 25, 2016, 2:30 p.m....institutions, or institutions they're thinking about applying to, are doing - for example, your benefits may...get rid of postdocs; that current postdoctoral appointments may shorten in length; that postdoc salaries...trainee workforce has clearly not been a sustainable approach to biomedical research in the U.S. and postdoctoral... -
The Challenges of Cell Culture
TypeBlog PostPublishedMay 5, 2016, 2:30 p.m....especially with industry. Today we also have the appropriate tools to verify our cells lines. Even if you ...the results are reliable. With this two-pronged approach, the many mistakes of the past can be prevented...including GSK, AstraZeneca-MedImmune, and Infors & Applikon. Due to his high level of experience he now travels... -
Harnessing Bacterial Toxins for Allelic Exchange
TypeBlog PostPublishedAug. 15, 2019, 12:30 p.m....References Espah Borujeni, Amin, Anirudh S. Channarasappa, and Howard M. Salis. "Translation rate is controlled...Scarless Modification of Proteobacterial Genomes." Applied and environmental microbiology (2019): AEM-00990...chromosomal gene replacement in Escherichia coli." Appl. Environ. Microbiol. 81.15 (2015): 5103-5114. PubMed... -
Fluorescent Proteins 101: GFP Fusion Proteins - Making the Right Connection
TypeBlog PostPublishedApril 9, 2019, 1:13 p.m....study its function is one of the most popular applications of fluorescent proteins. These fusion proteins...targeted to the endoplasmid reticulum based on the appearance of OSER whorls. This assay is quantitative and...linker sequence short In the early days of GFP applications, many were concerned with steric hindrance by... -
CRISPR 101: Validating Your Genome Edit
TypeBlog PostPublishedNov. 3, 2022, 12:15 p.m.... explain how to verify that your cells were appropriately edited for your desired mutation - insertion...frequency is 50%, then you could infer that approximately 25% of cells are likely -/-, 50% are -/+, and...sequencing confirmation. All knock-ins A method applicable to knock-ins both large and small is a software... -
New Optogenetic Tools for Cytoskeleton and Membrane Control
TypeBlog PostPublishedSept. 28, 2023, 1:15 p.m.... barrier of the endothelium, but they could be applied to study other actin-dependent processes in cells...) and to introduce unnatural head groups for applications like fluorescent labeling via click chemistry...constructs are great for in vitro biocatalytic applications where toxicity doesn’t matter. Plus, for folks... -
The Fluorescent Vegetables in Aptamer Soup
TypeBlog PostPublishedJan. 29, 2019, 2:27 p.m....enjoy a warm meal on a winter day, you should be happy to learn that many DNA-based reagents are named ...aptamer soup by exploring the development and applications of fluorescent RNA aptamers in Addgene’s collection...fluorescence when bound to DFHBI despite being approximately half the length (Warner et al., 2014); its length... -
Multiple Plasmids at a Low Price: Kits from Addgene
TypeBlog PostPublishedSept. 14, 2016, 2:30 p.m....polycistronic E. coli expression constructs. Some applications of plasmids from this kit include the production...can be engineered to bind specific RNA bases. By swapping out these different modular repeat segments, you...variety of functions via different fusions. Some applications of PUF protein fusions reported in the literature... -
CRISPR 101: Epigenetics and Editing the Epigenome
TypeBlog PostPublishedJune 24, 2020, 5:45 p.m....to specific DNA sequences. Non-editing CRISPR applications direct catalytically dead dCas9 fused to a variety... publication at Addgene. Klann TS, Black JB, Chellappan M, Safi A, Song L, Hilton IB, Crawford GE, Reddy...Ruan K, Wang F, Xu G, Hu R (2016) A CRISPR-based approach for targeted DNA demethylation. Cell Discovery... -
Hot Plasmids: Spring 2025
TypeBlog PostPublishedJune 3, 2025, 1:15 p.m....popular choice for a variety of synaptic imaging applications. Find iGluSnFR4 plasmids here! Aggarwal, A....that they could reprogram the genomic target by swapping the homology arms for new targets. And, importantly... is scarless, unlike previous integrase-based approaches. Find STITCHR plasmids here! Fell, C. W., Villiger... -
Viral Vectors 101: Optogenetic Tools
TypeBlog PostPublishedJune 27, 2023, 1:15 p.m....tools Once you’ve selected the class of tool appropriate for your experiment, you’ll need to find a specific...Yizhar, O., & Deisseroth, K. (2012). Principles for applying optogenetic tools derived from direct comparative... -
Technologies Enabled by NanoLuc® Luciferase
TypeBlog PostPublishedFeb. 8, 2018, 12:17 p.m....et al (2). In this post, I’ll cover two great applications of the NLuc technology. The first couples NLuc...the coupled luciferase. This strategy was first applied using RLuc8, an engineered Renilla luciferase, ...fluorescence. The paper reports a wide variety of applications of the Enhanced Nano-Lanterns including monitoring... -
Plasmids 101: Repressible Promoters
TypeBlog PostPublishedDec. 20, 2022, 2:15 p.m....for sphingomonads and other Alphaproteobacteria. Appl Environ Microbiol. 2013 Nov;79(21):6795-802. doi...cumate-inducible gene expression system for Bacillus. Appl Microbiol Biotechnol. 2019 Jan;103(1):303-313. doi...expression of genes for metabolic engineering applications in Saccharomyces cerevisiae.” FEMS Yeast Res... -
Transferable Skills Guide: Teamwork
TypeBlog PostPublishedNov. 28, 2017, 2 p.m....cross-team guy” at Addgene, an unofficial title that I happily embrace (and admittedly, I may have started that...identify as one. But some time around 10th grade, I snapped out of the phase of my life that had me listening...you’re part of a team - it will just make you a happier person. What does a good team need? Besides that... -
Quick Guide to All Things Lentivirus
TypeBlog PostPublishedMarch 21, 2017, 2:30 p.m....have been increasingly popular in fundamental and applied research since their first use in the early 90’... at transferring the gene of interest to the appropriate cells, it was obvious that its safety was a disadvantage...either tTA or rtTA (FUW-M2rtTA). In this kind of approach, a population of cells might express only the ... -
Starter Guide to induced Pluripotent Stem Cells (iPSCs) Part 2: Reprogramming and Transdifferentiation
TypeBlog PostPublishedNov. 6, 2018, 1:12 p.m....process is favored over the other. Cell conversion happens via: ReprogrammingA differentiated somatic cell...cells into another cell type. However, these two approaches differ in several ways. Below is a table listing...: 26042384. 27. Nejadnik, H., et al., Improved approach for chondrogenic differentiation of human induced... -
CRISPR 101: Multiplex Expression of gRNAs
TypeBlog PostPublishedSept. 10, 2020, 11:45 a.m....chance that all the edits you want to make will happen. There are many reasons to multiplex gRNAs, some...adjacent to the sites. When digested with the appropriate Type IIS enzyme, the unique flanking overhang...subsequent “bricks” to be added one by one. This approach is especially useful for combinatorial analyses... -
CRISPR 101: Cytosine Transversion Editors
TypeBlog PostPublishedMarch 25, 2025, 1:15 p.m....huge innovation, there was a limitation to this approach. CBEs and ABEs are base transition editors. They...GBEs), Xueli Zhang’s lab, returned with a unique approach to improving C → G editing. Instead of fusing ...abasic site. In this post, it refers to the same approach as a CGBE. BER Base Excision Repair A cellular... -
New Antibodies in the Lab Animation!
TypeBlog PostPublishedNov. 19, 2024, 2:15 p.m....Thinking about using antibody-based applications in your experiment...but not sure where to start or ...or quick refresher on — these common antibody applications. Thanks for watching, and we hope you found... -
#DeckTheLab to Win a Tasty Winter Treat This Holiday Season
TypeBlog PostPublishedNov. 30, 2015, 3:30 p.m....wintertime festivities! Help us spread some cheer by snapping a photo of you and your labmates celebrating the... the New Year. Will your lab be a lucky winner? Happy holidays from all of us at Addgene! We look forward... -
'Tis the Season to #DeckTheLab
TypeBlog PostPublishedDec. 2, 2016, 3:30 p.m.... Why #DecktheLab? Help us spread some cheer by snapping a photo of you and your labmates celebrating the... the New Year. Will your lab be a lucky winner? Happy holidays from all of us at Addgene! We look forward...