...with Gibson assembly. While this is a comprehensive mutagenesis method, a drawback to using Gibson assembly...plasmids ready for deep mutational scanning.     3 reasons to be interested in deep mutational scanning Hundreds...rate of mutagenesis, and low throughput.       Gibson Assembly: An easy-to-use technique that joins homologous...especially single basepair deletions. Therefore, Gibson assembly is better suited for creating small, focused... PubMed PMID: 27723752.  4. Fowler, Douglas M., Jason J. Stephany, and Stanley Fields. “Measuring the ...

...2015b) under a CC-BY 4.0 license. Long et al., Nelson et al., and Tabebordbar et al. each chose to try.... used SpCas9 in conjunction with AAV9, whereas Nelson et al. and Tabebordbar et al. used the shorter ...not subject to nonsense-mediated decay. Although Nelson et al. observed only 2% genome editing in one experiment...persist long-term. In accordance with this result, Nelson et al. find that dystrophin restoration is maintained... E, Bhattacharyya S, Shelton JM, Bassel-Duby R, Olson EN. Postnatal genome editing partially restores ...Dec 31. pii: aad5725. PubMed PMID: 26721683. 3. Nelson CE, Hakim CH, Ousterout DG, Thakore PI, Moreb EA...

...path. Because there is no demand for a favor, the person you ask will be more likely to say yes and more... Informational Interviews are best conducted in person. Perhaps you can offer to buy coffee or lunch at... your interviewee? It is always best to meet in person to develop relationships, but if it is not possible.... Keep in mind that this is a good time to ask personal questions about salary, opportunities for advancement...advancement, challenges and disappointments. These more personal questions can elicit the most valuable insights...

...secrets - during a collaboration, members of Maureen Hanson’s and Rima Menassa's labs made the accidental discovery...created a transgenic GFP-containing organism,” says Hanson, “using a gene encoding one of the unexpectedly...Sattarzadeh A, Saberianfar R, Zipfel WR, Menassa R, Hanson MR. Green to red photoconversion of GFP for protein... PubMed. McKinney SA, Murphy CS, Hazelwood KL, Davidson MW, Looger LL. A bright and photostable photoconvertible...from this publication at Addgene. Subach OM, Patterson GH, Ting LM, Wang Y, Condeelis JS, Verkhusha VV...

... should I try research? Personally, I have always been a pretty curious person. I like to ask questions...anyone else in the world knows it—to be the first person to make a discovery. I can’t tell anyone else what...place to begin. Here’s my advice: Reflect on your personal interests While it’s perfectly fine to be unsure... also makes for an experience more tailored to personal interests. Talk to people, ask questions, and ...First-Generation College Students. Social Psychological and Personality Science 11:647–657 . https://doi.org/10.1177/...

...DNA. Finally, in 1953, Franklin along with James Watson and Francis Crick resolved the DNA double helix...engineering itself is attributed to SciFi writer Jack Williamson who used it in his 1951 novel Dragon’s Island...movies. Rampage (2018), starring Dwayne “The Rock” Johnson, calls out CRISPR as the genetic engineering experiment...Davis JR, Sousa AA, Koblan LW, Levy JM, Chen PJ, Wilson C, Newby GA, Raguram A, Liu DR (2019) Search-and-replace...Gaudelli NM, Komor AC, Rees HA, Packer MS, Badran AH, Bryson DI, Liu DR (2018) Publisher Correction: Programmable...:3870–3879 . https://doi.org/10.1093/nar/gkf495 Watson JD, Crick FHC (2003) Reprint: Molecular Structure...

...and Hoess, 1984; Van Duyne, 2001; Christenson Wick and Krook-Magnuson, 2018). Optogenetic FLEx vectors ...PubMed PMID: 11340053. Wick, Zoé Christenson, and Esther Krook-Magnuson. "Specificity, versatility, and...Nature 351.6326 (1991): 489. Hirsch, Matthew L., Sonya J. Wolf, and R. J. Samulski. "Delivering transgenic...PMCID: PMC5111696. Towne, Chris, and Kimberly R. Thompson. "Overview on research and clinical applications...

...technological change happens really fast,” Thompson says. To do this, Thompson reached out to MIT labs in the Synthetic... classification This inquiry started when Neil Thompson, a research scientist at MIT’s Computer Science...query coverage) that could be determined through comparisons with a reference sequence database. Through a...Kunjapur, Aditya M., Philipp Pfingstag, and Neil C. Thompson. "Gene synthesis allows biologists to source genes...

...proteins without changing excitation wavelength (Salomonnson et al. 2012). They can also be excited efficiently...emission intensities from fluorescent proteins (Salomonnson et al. 2012). Finally, it is worthwhile to test...18718768. PubMed Central PMCID: PMC2569002. 3. Egen, Jackson G., et al. "Macrophage and T cell dynamics during...18261937. PubMed Central PMCID: PMC2390753. 4. Egen, Jackson G., et al. "Intravital imaging reveals limited ...21527931. PubMed Central PMCID: PMC4772972. 11. Salomonnson, Emma, et al. "Cell-based and in vivo spectral...Wolfgang Weninger. "Dendritic cell behaviour in vivo: lessons learned from intravital two-photon microscopy."...

...favourable ratings to the abstracts that affirmed their personal beliefs even though all three texts were otherwise...were exhibiting a phenomenon called “motivated reasoning,” the catchall term for a range of neat little...little tricks our minds perform to avoid cognitive dissonance (Kunda, 1990). More specifically, they were engaging...sources of information they don’t like, looking for reasons to dismiss them. There’s evidence to show that ...nclimate1547 Kunda, Z. (1990). The case for motivated reasoning. Psychological bulletin, 108(3), 480. More resources...

...effectively contribute as part of a team.” As a seasoned educator, Kuldell also knew that the best days...student engagement and joy, rather than a perfect lesson plan or outstanding test scores.  As a result, ...are provided with synthetic biology lab kits and lesson plans adapted from her own MIT curriculum. Drawing...investigate together. Teachers decide which BioBuilder lessons to teach, and how — some choose to run full semester-long...semester-long classes, some pick and choose a few lessons to include each year; others use BioBuilder as ...

...essential for the structure of paraspeckles. Clemson CM, Hutchinson JN, Sara SA, Ensminger AW, Fox AH, Chess...your favorite biological phenomenon, there’s a reasonable good chance that Addgene can help. Though it ...Fox Elaine Fuchs Kai Ge Robert Kingston Thoru Pederson Phil Sharp Julie Ahringer Alexander Stark Bas..., especially Karen Reddy (@theReddyLab), Thoru Pederson, Joanna Wysocka, Kerstin Bystricky, Ben Martin...

..., and substitutions - at the same efficiency as Gibson Assembly and GeneArt cloning - but at a much lower...digested and ligated into an appropriate vector. Gibson Assembly, GeneArt Cloning, and SLIC can all be ...When REPLACR-mutagenesis goes head-to-head with Gibson assembly and GeneArt cloning, the results are favorable...hands!     REPLACR-Mutagenesis Megaprimer PCR Gibson Assembly GeneArt Cloning Types of Mutations Insertions...Made Competent Cells Required Yes No No No Comparison of REPLACR-mutagenesis with other common methods...

...Twitter, LinkedIn and Facebook to see their online personas. Some have online coaching tools that can be very...remotely (by phone, Skype or Google Hangout) but in-person coaching is nice if you can get it.  Because I ...this offers the option of matching with the best person in the group for your needs. Beth Schachter of ... Michelle Capes at Adeptify Me. While based in Madison, Wisconsin she also works remotely and has experience...support for issues of diversity in science.  A great person to contact is Twitter’s @DrQualls. Marquita Qualls...

...genetic tools. Mobile-CRISPRi from the labs of Jason Peters, Oren Rosenberg, and Carol Gross provides...Mobile-CRISPRi is introduced using the bacterial Tn7 transposon system. This system has two parts: a plasmid ...Introducing Mobile-CRISPRi into bacteria. Top: A Tn7 transposon containing the CRISPRi components and a plasmid...methods 2.6 (2005): 443. PubMed PMID: 15908923. Johnson, Christopher M., and Alan D. Grossman. "Integrative...4.2 (2019): 212. PubMed PMID: 30675034. Peters, Jason M., et al. "Enabling genetic analysis of diverse...

... Jake Watson and Javier García-Nafría from the MRC Laboratory of Molecular Biology. Jake Watson is a postdoctoral...This post was contributed by Jake Watson and Javier García-Nafría from the MRC Laboratory of Molecular...receptor function.     Follow them on Twitter @jakefwatson and @JGarciaNafria.  References 1. Jones, Douglas...PMCID: PMC331480. 3. García-Nafría, Javier, Jake F. Watson, and Ingo H. Greger. "IVA cloning: a single-tube...

...libraries that do just that (Doench et al., 2016, Sanson et al., 2018). These libraries utilize optimized...additional transcription factors to the targeted region (Sanson et al., 2018) . This tracrRNA was combined with...selected per gene and divided into two sets, A and B. Sanson et al. demonstrated that the Calabrese library ... more hits than the SAM library approach in a comparison study screening for vemurafenib resistance in...: 25494202. PubMed Centra PMCID: MC4420636.  5. Sanson, Kendall R., et al. "Up, down, and out: next generation...

...Kostick, K., Torgerson, L., Muñoz, K. A., Hsu, R., Kalwani, L., Sierra-Mercado, D., Robinson, J. O., Outram... rat CNS: AAV PHP.EB vector dose–response and comparison to AAV PHP.B. Gene Therapy, 25(5), 392–400. https...00981-0 Wright, J. F., Le, T., Prado, J., Bahr-Davidson, J., Smith, P. H., Zhen, Z., Sommer, J. M., Pierce... Jason Nasse...

... to put to rest a common misconception. I find personal bias often unintentionally plays a big role in...varied in how they prefer to consume information. Personally, I would much rather read the instructions than... accessible via web searches; likewise, most in-person events and venues will have a solid understanding...Primarily useful for data-focused stories. In-person lectures, presentations, etc… Low (except for ...  Rachel Leeson...

...As a person with many roles and responsibilities, I have to think a lot about how to balance those roles...your community’s schools, here are a few tips and lessons I’ve picked up from my recent experiences. STEM...history helped demonstrate that a) I was a real person who used to be a middle schooler himself and b)...back and trying again next year. There are many reasons why you might want to reach out to your school ...tumultuous time in scientific history. Whatever your reasons, let us know how it goes! Acknowledgements: I...

...advice, open doors for you, and teach you valuable lessons along the way. We think it’s so important that ... technician, or a graduate student. This is the person you report or talk to on a daily basis who will...scared to ask for some! You can email or ask in-person “Are there any papers you think would be useful...these things will help you get more out of the in-person training component. Documenting during training...take on trainees one day when you are the senior personnel! Happy training! Resources Resources on the Addgene...

...CAC trinucleotides during neuronal maturation (Tillotson & Bird, 2019) and is a global transcriptional ...rescue the neurological defects of Rett syndrome (Tillotson et al., 2017). Advancing gene therapies for Rett...Supplement/S51.002.abstract Sinnamon JR, Kim SY, Fisk JR, Song Z, Nakai H, Jeng S, McWeeney SK, Mandel G (2020)... . https://doi.org/10.1016/j.omtm.2017.04.006 Tillotson R, Selfridge J, Koerner MV, Gadalla KKE, Guy J...398–401 . https://doi.org/10.1038/nature24058 Tillotson R, Bird A (2020) The Molecular Basis of MeCP2 ...

...service in 2016. We realized we could apply the lessons learned from our plasmid lab to develop a reliable...available. This prioritization happens for good reasons. We began offering our viral vector service because...our Viral Vectors 101 eBook and blog posts. The lessons learned from this process have shaped and supported...fnana.2019.00093 McDade, J. R., Waxmonsky, N. C., Swanson, L. E., & Fan, M. (2016). Practical Considerations... Rachel Leeson...

...are addressed with ddPCR technology (Hindson et al., 2011, Hindson et al., 2013), which has been shown ...2011): 8604-8610.4. Hindson CM, Chevillet JR, Briggs HA, Gallichotte EN, Ruf IK, Hindson BJ, Vessella RL,... therapy 6.7 (1999). PubMed PMID: 10455443. 3. Hindson, Benjamin J., et al. "High-throughput droplet digital...

...organisms have different codon usage biases? The reasons for varied codon preferences among organisms aren...aren’t completely understood, but some possible reasons include: Metabolic pressures - it takes cellular...recognize codons found in highly expressed genes (Emilsson and Kurland, 1990). Controlling gene expression...15678157. PubMed Central PMCID: PMC1299251. 3. Emilsson, Valur, and Charles G. Kurland. "Growth rate dependence... 2265611. PubMed Central PMCID: PMC552224. 4. Gustafsson, Claes, Sridhar Govindarajan, and Jeremy Minshull...

...Pinnelli M, Ansari R, Harper S, Jackson DA, McRae R, Pooley R, Wilkinson P, van der Meer D, Dow D, Buser-Doepner...of TSS for best results but up to 400 bp works reasonably well.  Best of luck on your experiments!  Many...Housden BE, Perrimon N, Collins JJ, Church G (2016) Comparison of Cas9 activators in multiple species. Nat Methods...doi.org/10.1038/nmeth.3871 Gilbert LA, Horlbeck MA, Adamson B, Villalta JE, Chen Y, Whitehead EH, Guimaraes...

.... 4. Shaner, Nathan C., George H. Patterson, and Michael W. Davidson. "Advances in fluorescent protein... structure resembles images of the deity (see comparison image above). The church of the Flying Spaghetti...schools and was started in 2005 in Kansas by Bobby Henderson, a physics graduate from Oregon State University...New York. pp 719. 3.Rizzo, Mark A., Michael W. Davidson, and David W. Piston. "Fluorescent protein tracking...

...know your collaborators from other institutions personally on conference calls and make sure to maintain...collaborating with people outside my lab’s department, I personally felt that I had to overcome some hesitation to...the hard part: maintaining these relationships. Personally, I find this to be the most challenging and time-intensive...maintain a relationship - many of them involve personal meetings and common interest so naturally this...sharing. LinkedIn or ResearchGate allow you to send personal updates as messages and keep you up-to-date on...

...favorably to other methods. In a head-to-head comparison with Cas9, Cas9 modified 9/12 known off-targets... C., Rees, H. A., Packer, M. S., Badran, A. H., Bryson, D. I., & Liu, D. R. (2017). Programmable base ...doi.org/10.1038/nbt.3803 Koblan, L. W., Doman, J. L., Wilson, C., Levy, J. M., Tay, T., Newby, G. A., Maianti... Bohnuud, T., Fernandez, T. P., Gehrke, J. M., Grayson, P., Lee, K. D., Ortega, M. A., Sawyer, C., Schwaegerle..., Lapinaite, A., Newby, G. A., Thuronyi, B. W., Wilson, C., Koblan, L. W., Zeng, J., Bauer, D. E., Doudna...Levy, J. M., Yeh, W.-H., Zheng, C., Newby, G. A., Wilson, C., Bhaumik, M., Shubina-Oleinik, O., Holt, J....

...10.3390/antib8020028  Bird R, Hardman K, Jacobson J, Johnson S, Kaufman B, Lee S, Lee T, Pope S, Riordan... (Sandomenico et al., 2020). Figure 1: Comparison between the IgG antibody and scFv.   What...doi.org/10.1371/journal.pone.0189964  Huston JS, Levinson D, Mudgett-Hunter M, Tai MS, Novotny J, Margolies... made by molecular evolution 1 1Edited by I. A. Wilson. Journal of Molecular Biology 275:245–253 . https...

...cells are great model organism not only for the reasons above but because they actually share many biological... the study of neurological diseases, such as Parkinson’s disease (PD). PD is primarily characterized by...pancreas and produce different levels of insulin (Hesselson et al., 2009). The worm (Caenorhabditis elegans...nature391.6669 (1998): 806. PubMed PMID: 9486653. Hesselson, Daniel, et al. "Distinct populations of quiescent...From the baker to the bedside: yeast models of Parkinson's disease." Microbial cell 2.8 (2015): 262. PubMed...

...designing and testing your AAV.  Figure 2: Comparison of AAV serotypes 1, 2, 5, 6, 8, and 9 in two ...Taschenberger et al., 2017).     Figure 3: Comparison of two astrocyte selective promoters demonstrating...Sortwell, C. E., Hauswirth, W. W., Boye, S. E., & Manfredsson, F. P. (2017). Rationally Engineered AAV Capsids...vector-mediated transduction of the corticospinal tract: Comparison of four promoters. Gene Therapy, 28(1–2), 56–... Jason Nasse...

...PMC2693335. Williams JA, Luke J, Langtry S, Anderson S, Hodgson CP, Carnes AE. Generic plasmid DNA production...and at high copy numbers (Williams, Carnes, and Hodgson, 2009). More research is ongoing to determine plasmid...90057-7. PMID: 2544913. Cook KS, Luo J, Guttman A, Thompson L. Vaccine Plasmid Topology Monitoring by Capillary...90060-6. PMID: 6323019. Williams JA, Carnes AE, Hodgson CP. Plasmid DNA vaccine vector design: impact on...

...2013), or rescue of mutant phenotypes (unc-119) (Dickinson et al., 2013). These approaches allowed effective...the development of the self-excising cassette (Dickinson et al., 2015) made this step as simple as heat-shocking...e193–e193 . https://doi.org/10.1093/nar/gkt805 Dickinson DJ, Pani AM, Heppert JK, Higgins CD, Goldstein...sequences https://doi.org/10.17912/W2237D Farboud B, Severson AF, Meyer BJ (2018) Strategies for Efficient Genome...Krishnamurthy PM, Gneid H, Edraki A, Watts JK, Sontheimer EJ, Mello CC (2018) 5′ Modifications Improve ...

...many stages of synthesis and degradation. For this reason it is virtually impossible to determine specific...parameter to consider is the contrast ratio- a comparison of PA-FP brightness before and after photoactivation...extraordinary molecular detail.   References 1.Patterson, George H., and Jennifer Lippincott-Schwartz. ..., Samuel T., Thanu PK Girirajan, and Michael D. Mason. "Ultra-high resolution imaging by fluorescence ...Photoactivatable FPs in Empty Backbones in the Michael Davidson Collection Find Photoactivatable FPs for Single...

...contributed by Kurt Swanson, husband of Addgene Director of Biology, Lianna Swanson, and researcher at ...in food products). I picked this post for a few reasons: 1) I think phage are super cool and have considered...proteins that could be useful for a wide variety of reasons. 3) I would love to see more posts like this on...

... spend a few days editing and later looking for songs and sounds to synchronize with the images thereby...the hardest part of the process was to embed personality in pieces of plastic, but it’s gotten easier ...than a piece of plastic for us now). With the personality in place, everything now is about the plot. This...How You Can Use Video to Share Your Research 5 Reasons to Use Reddit for Science Communication 10 Ways...

...to improve, and they’re also clear about how the person is doing overall. Employees hate to wonder what...employee. Make sure to focus on the action, not the person - intent is usually good. Don’t pretend something...standard format not only enables you to discuss a person's performance or what has been learned, it helps... effective over time if you take it seriously.  Person who will receive feedback does a self-assessment...

... ask a lot of questions. Personality and style conflicts are a common reason for scientists to change ...budgeting, equipment acquisition, public speaking and personnel management?  If you are thinking about a non-academic... you can help it. Observe as much as you can in person. Rotate in the lab, if possible. Attend a lab meeting...

...use Bioluminescence Resonance Energy Transfer (BRET) were developed by the Johnson Lab . Characteristics...CRISPR labeling chromosomal loci in genome Thoru Pederson’s lab at the University of Massachusetts Medical...lab further modified sdChr- a blue-shifted channelrhodpson from freshwater green alga identified by Ed...Nano-lanterns rely on the principle of bioluminescence resonance energy transfer (BRET), whereby photons emitted...-tagged human ORF clones to allow testing and comparison of different cell-free expression systems; the...understanding complex gene regulation (Bert et al 2000, Johnson et al 2004). So if you are looking for a luciferase... et al. Plasmid. 2000. PubMed PMID: 10964627 Johnson, et al. Mol Cell Biol. 2004. PubMed PMID: 15340054...