We narrowed to 640 results for: SON;

...modify a given gene. CRISPR-tranposons By combining CRISPR editing and transposons, the Sternberg and Zhang...pneumoniae. Robert Husson lab's M. tuberculosis CRISPRi plasmids A group from the Husson lab used CRISPRi...insert extra elements, like a repair template, using Gibson Assembly or restriction enzyme cloning. Streptomyces...Zhang labs developed CRISPR-transposons. The Sternberg lab's system called INTEGRATE (Insertion of Transposable...By using a multi-spacer CRISPR array, CRISPR-transposons can be multiplexed. Transcriptional repression...CRISPRi for Gram-negative and Gram-positive bacteria Jason Peters, Oren Rosenberg, and Carol Gross used two...https://doi.org/10.1038/s41564-018-0327-z Qi LS, Larson MH, Gilbert LA, Doudna JA, Weissman JS, Arkin AP...

...compatible cohesive ends Engler et al., 2008 Gibson Cloning Gibson assembly method Single reaction, can assemble...Central PMCID: PMC2574415. 3. Gibson DG, Young L, Chuang RY, Venter JC, Hutchison CA, Smith HO.Enzymatic assembly...terminus or the 3’ terminus of the ligation product. Comparison to other high throughput cloning strategies There... identical polyadenylation signals or promoters Gibson et al., 2009 MXS-Chaining, BioBricks, Bglbricks...Sladitschek and Neveu, 2015; Shetty et al., 2008; Anderson et al., 2010 Among the chaining-based methods...Additional Resources on the Addgene Blog Read about Gibson Assembly Learn About Other Plasmid Cloning Techniques... Find Plasmid Cloning Protocols References 1. Anderson JC, Dueber JE, Leguia M, Wu GC, Goler JA, Arkin...

..., based on resorufin). SNAP-tag: In 2003, Kai Johnsson’s lab developed a self-labeling fusion tag based...Addgene’s collection. CLIP-tag: Also in 2008, Kai Johnsson’s lab reported another AGT-based tag named CLIP-tag...STED) nanoscopy. Scale bar, 2 μm. Inset shows comparison with standard confocal microscopy. Image adapted...159 aa) are smaller than HaloTag (295 aa) – for comparison, common fluorescent proteins are typically 230...Wissner, R. F., Xi, Z., Allgeyer, E. S., Zhong, S., Thompson, A. D., Lowe, N., Butler, R., Bewersdorf, J., ...Strategies Matter for Super-Resolution Microscopy: A Comparison between HaloTags and SNAP-tags. Cell Chemical...Corrêa, I. R., Kindermann, M., Beaufils, F., & Johnsson, K. (2008). An Engineered Protein Tag for Multiprotein...

... you want to make will happen.  There are many reasons to multiplex gRNAs, some being: Using dual nickases...the Bullock lab, and gRNAs can be inserted using Gibson Assembly or SLIC cloning methods. A BsaI-based ... up your multiplexed gRNAs with Golden Gate and Gibson assembly methods If you want to scale up the number...use some assembly methods such as Golden Gate or Gibson assembly. Golden Gate Assembly of gRNAs Golden ...supply it with another plasmid.   Figure 3: Comparison of Multiplex Strategies including Standard PolIII-gRNA...Multiplexing in plants Qi-Jun Chen Lab Golden Gate/Gibson Assembly Multiplexing Plasmids: These plasmids ...you to assemble 2-4 gRNAs through Golden Gate or Gibson Assembly. gRNAs are inserted into the pCBC vectors...

...process known as opsonization. In antibody-dependent complement deposition (ADCD), opsonization of IgG and ...isotype class or even subclass, and are part of the reason why different isotypes drive different types of... in the pathogen’s cell membrane (Dunkelberger & Song, 2010). As cells cannot survive when holes are punched...(ADCC) is a slightly more direct method than opsonization. In ADCC, IgG antibodies bind to a pathogen ..., our antibodies have induced “stabbing” and “poisoning,” but let’s be honest, the elimination style the...10.1038/74704. PMID: 10742152. Dunkelberger, J., Song, WC. Complement and its role in innate and adaptive...  Rachel Leeson...

...making the detection of rare events difficult. Dickinson et al. sought to create a new and efficient selection-based...Figure 1: Self-excising cassette schematic from Dickinson et al. (2015). Used with permission from the Genetics...your gene of interest into the FP-SEC vector using Gibson assembly. This construct will serve as the repair...for the targeted gene. After initial testing, Dickinson et al. tested this system across 7 genes in parallel...modification. Figure 2: Schematic from Dickinson et al. (2015) illustrating the steps to fluorescently...Society of America. Applications of this system Dickinson et al. have developed a modular system that requires...with a Self-Excising Drug Selection Cassette. Dickinson DJ, Pani AM, Heppert JK, Higgins CD, Goldstein...

...background information. Your options for this step are sonication, which uses high-frequency sound waves to break...break up the DNA, or enzymatic digestion. Sonication provides a truly random digestion, but is harsher ...to the selective nature of the cutting. While sonication is usually the preferred method, enzymatic digestion...concentration.  Protein Capture After crosslinking and sonication or enzymatic digestion, you’re ready to incubate...availability, which could affect antibody selection. Sonication (or enzymatic digest) will also have to be optimized...fragment length for your output assay. A longer sonication time could mean needing to increase crosslinking...effects of increased sample agitation; a shorter sonication time could be an opportunity to decrease crosslinking...

...PubMed Central PMCID: PMC146043. 9. Nilsson, Stefan V., and Göran Magnusson. "Sealing of gaps in duplex DNA...gaps. For example, DNA assembly methods, such as Gibson Assembly® and NEBuilder® HiFi DNA Assembly, require...for a ligase? DNA ligases generally prefer fully Watson-Crick base-paired dsDNA substrates to those containing... base-pair size nearly indistinguishable from a Watson-Crick base pair, are joined with nearly the same...phosphate to the ligation). The structural/mechanistic reason for this differential is not known for certain,...allowed screening of all possible base combinations (Watson-Crick and mismatched) around the ligation junction....6 (2004): 757-764. PubMed PMID: 15582400. 2. Tomkinson, Alan E., et al. "DNA ligases: structure, reaction...

...I first became a manager. Know your people Each person is unique and should be managed accordingly.  I...role is not to do all the work yourself anymore. Personally I found this to be particularly challenging. ...important” tasks my schedule. This was bad for 2 reasons. First, it soon became obvious that I didn’t have...accordingly -  but think of it as an investment: once the person is trained you will have more time to focus on ...identify what to delegate and to whom (make sure that person has time for it!). Then schedule a meeting to discuss...Unless you’re delegating a recurring task to a seasoned report, there should be questions! Remember that...learned to embrace them after one ended with the person in front of me saying “I’ve been doing some of ...

...the Deverman lab at the Broad Institute and the Wilson lab at the University of Pennsylvania independently...unlikely receptor for PHP.B, and how the Deverman and Wilson labs determined LY6A was a receptor used by PHP.B... PHP.B CNS permissiveness Both the Deverman and Wilson labs determined LY6A, also known as stem cell antigen... role in viral susceptibility. The Deverman and Wilson Labs independently identified LY6A as a receptor...was a likely player in PHP.B permissiveness. The Wilson Lab also looked for the gene variant responsible...require galactose and AAVR to transduce cells. The Wilson Lab generated results similar to the Deverman Lab...’s transduction of 293T cells. In addition, the Wilson Lab also tested the ability of PHP.B to transduce...

... of your lab that you see in person, and helps improve your interpersonal relationships with people you...emailing documents to one person at a time. Just be sure there is a point person responsible for incorporating...replicates how you’d interact with team members in person, and improves overall communication.  Maintain...out if any milestones are missed. Spend time in person Presenting the results from my lab exchange...having collaborators spend time with each other in person. One possibility is to do a lab exchange, where...

...to our guest blogger Lydia Morrison from New England Biolabs. Lydia Morrison is a biochemist by training...This post was contributed by guest blogger Lydia Morrison from New England Biolabs. What is DNA assembly...multiple methods of DNA assembly available, including: Gibson Assembly®, BioBrick® Assembly, Golden Gate Assembly...Assembly, and NEBuilder® HiFi DNA Assembly. Gibson Assembly allows the production of scarless DNA constructs...opens the door for new opportunities in the field.  Gibson Assembly® is a registered trademark of Synthetic... Addgene Blog Learn More about Gateway Cloning Gibson Assembly and Other Long-Homology Based Cloning ...

...This post was contributed by guest blogger Chris Richardson, a Postdoctoral Researcher in Jacob Corn’s lab...and yet unanswered) question. Our recent paper (Richardson et al 2016) revealed some of the biophysical ...binding to the cleaved DNA duplex. Image from Richardson et al., 2016. We next wondered if we could...interaction with the Cas9-DNA duplex. Image from Richardson et al., 2016.   dCas9 enables sequence replacement...Results from our lab and others (Lin et al 2014, Richardson et al 2016, and Kim et al 2014) have shown that...transcribed sgRNA, and optional donor DNA. Chris Richardson is a Postdoctoral Researcher in Jacob Corn’s ...worked on this publication:         References 1. Richardson, Christopher D., et al. "Enhancing homology-directed...

...called GMAP (Gibson assembly-based modular assembly platform). GMAP is based on the Gibson Assembly method...Parton DL, Hanson SM, Rodríguez-Laureano L, Albanese SK, Gradia S, Jeans C, Seeliger MA, Levinson NM, Chodera...as individual plasmids shipped as agar stabs. Iverson, et al. ACS Synth Bio. 2015. PubMed PMID: 26479688...30-40bp overlaps). To expand the capabilities of Gibson assembly and make a more modular platform, the ...context-dependent human fitness genes, the laboratory of Jason Moffat at the University of Toronto has created ...Vectors Available at Addgene: Lab Plasmids Dan Carlson RCIscript-GoldyTALEN, pC-GoldyTALEN Tom Ellis...pTAL7a, pTAL7b Michal Zurovec pBlue-TAL Nathan Lawson pJDS Series   Superglue proteins with SpyTag...

...is. If the person makes time for you, as needed, this is a strong indicator that this person cares about...limited job opportunities in academia - the main reason most people enter postdoctoral training. Similarly... the postdoctoral mentor, and at least one more person that knows you and your research well. Who could...and skilled speaker. There's a more fundamental reason to give presentations - networking! People get ...unpublished information and reagents. Part of the reason I got my first faculty position was because the...institute about me, even though I had not asked this person to write a letter of reference. The group leader...not take time off when you are a PF. Here is my personal list:   Exercise - nothing dissipates stress ...

...Johnny Kung, Director of New Initiatives for the Personal Genetics Education Project (pgEd). Advances in...exacerbating existing inequalities and injustices. The Personal Genetics Education Project (pgEd), where I currently...conversations about both the benefits as well as the personal, social and ethical implications of genetics. ...cultural or religious affiliation, and ethnic or personal identity. We do this through a variety of activities...activities, including creating educational materials and lesson plans that are made freely available online; running...translating all these into social media and blog posts, lesson plans, or information briefs for policymakers. ...Kung is the Director of New Initiatives for the Personal Genetics Education Project (pgEd). He is broadly...

...labelled with horseradish peroxidase (HRP; Kristensson and Olsson, 1971), plant lectins (wheat germ agglutinin...24336151. PubMed Central PMCID: PMC4004172. Kristensson, K., and Olsson, Y. (1971). Retrograde axonal transport...difficult to accurately map network connections (Lo & Anderson, 2011). In the past two decades, several virus...PubMed Central PMCID: PMC4324159. Castle, M.J., Gershenson, Z.T., Giles, A.R., Holzbaur, E.L., and Wolfe...29, 363–365. PubMed PMID: 4107258. Lo, L., and Anderson, D.J. (2011). A Cre-dependent, anterograde transsynaptic..., J.S., Hyman, B.T., and McLean, P.J. (2009). Comparison of transduction efficiency of recombinant AAV... V., Thiry, I., Deroose, C. M., Mortelmans, L., Wilson, J.M., Debyser, Z., and Baekelandt, V. (2007). ...

...a price of a few dollars per reaction. By comparison, Gibson assembly can cost over $15.00 per reaction...In today’s post, we will focus on a high-level comparison of popular cloning methodologies, with a special...as 30 minutes! Alternatively, you might consider Gibson cloning. In this technique, you first perform a...reaction if you purchase a pre-made mix. Nonetheless, Gibson assembly is a highly versatile and efficient way...propagating. The ligation-independent methods like Gibson and SLIC are also quite efficient; although efficiency...them are some of the homology based methods, like Gibson or SLIC, since one can pool the components and ...

...used to call copy numbers should be as small as reasonably possible and set using the variance seen within...negative for each reaction, math happens (this time, Poisson statistics), and the readout gives an absolute ... through concentration values, rather than by comparison to a standard curve, and most results can be ...7778-9_9. PMID: 29717442. Arya M, Shergill IS, Williamson M, Gommersall L, Arya N, Patel HR. Basic principles... S., Hollox, E. J., & Boon Peng, H. (2015). A comparison of assays for accurate copy number measurement...journal.pone.0116791 Providenti MA, O'Brien JM, Ewing RJ, Paterson ES, Smith ML. The copy-number of plasmids and ... Rachel Leeson...

...students who are unable to attend in-person interviews for a variety of reasons (logistical, financial, etc…)...visit in person, things like sickness or scheduling issues can and do interfere with those in-person plans...COVID-19 and standards for hybrid, virtual, and in-person events change seemingly daily (and are inconsistent...about the program and having the time to ask personalized questions might be difficult. However, an online...Even if you had planned a campus visit and the in-person portion got canceled, taking advantage of virtual...