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Showing: 281 - 300 of 560 results
  1. 5 Great Apps for Lab Life

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    ...misplacing your results or losing that lab notebook from 2 years ago. The free subscription to Lab Archives allots...notebooks you’re saving trees! The Earth thanks you. 2. Gibco Cell Culture by Gibco Whenever I start to learn... Lastly, the Tm calculator helps you set up an optimal PCR depending on the primer sequence and the NEB...
  2. Cancer, Inflammation and Immunity - Harnessing the Body’s Defenses to Fight Cancer

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    ...represent a favorable state for tumor progression (Ref 2). Future treatments may need to target this immunosuppressive...cell 30.1 (2016): 147-160. PubMed PMID: 27411589. 2. Todoric, Jelena, Laura Antonucci, and Michael Karin...starvation on cancer therapy. Findings in previous animal studies suggested that fasting has a positive effect...
  3. Plasmid-based Recombinant Monoclonal Antibodies: What They Are and Why You Should Be Excited About Them

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    ...use antibodies made from animals or hybridomas.   Figure 2: (Left) Figure 2: (Left) The light chain...Polyclonal antibodies produced in animals may be variable among animals and bleed dates and even clonal ...1: (Top) Polyclonal antibodies are produced in animals and consist of a mixture of antibodies recognizing... display, yeast display, ribosome display, or mammalian display (Tsuruta et al., 2017). In vitro selection...antibodies to targets that may not work well in animals because of similarities to host proteins. After...into host cells, such as bacterial, yeast, or mammalian cells, for antibody production and subsequent ...genetically drift over time. Antibodies made from animals are limited in supply and producing antibodies ...
  4. Recombinase-based State Machines Enable Order-dependent Logic in vivo

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    ...certain states of interest. For example, in Figure 2 below we present a two-input, five-state GRSM register...linking RSM inputs to endogenous gene expression (2). Our paper (1) explains how to arrange attB-attP...Science. 353, aad8559 (2016). PubMed PMID: 27463678. 2. Green, Alexander A., et al. "Toehold switches: de-novo-designed...its target states are only represented in a very small number of registers. Instead, one may use qPCR to... not be suitable for applications that require minimal cellular perturbation. Also, this method requires...
  5. 28 Hot Plasmid Technologies from 2015

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    ... Zetsche et al., Nat Biotechnol. 2015 Feb 2;33(2):139-42. doi: 10.1038/nbt.3149. Light-inducible ...and on your favourite signalling pathways (Figure 2). They are all available now at Addgene. 1 Lungu...20;19(4):507-17. 10.1016/j.chembiol.2012.02.006. 2 Guntas et al., Proc Natl Acad Sci USA 2015 Jan 6; ...Nihongaki et al., Chemistry and Biology. 2015 Feb 19; 22(2):169-74. High efficiency cloning for enzyme screening...inducible, gene knockdown with shRNA or shRNA-miR30, (2) tamoxifen-inducible CreERT2 gene deletion, (3) simultaneous...different fluorescent proteins, 8 constitutive promoters, 2 includible promoters, 3 polyA terminators and various...of plasmids required for each input/output system; 2) removing all chemically inducible promoters; 3) improving...
  6. Plasmids 101: Terminators and PolyA signals

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    ...due to the presence of additional helper sequences (2-3). As alluded to above, termination and polyadenylation...and translation processes as well.   Figure 2: The conserved eukaryotic polyadenylation signal directs...types of RNA: Polymerase I is responsible for ribosomal RNA, Polymerase II is responsible for mRNA and...seems to couple termination with polyadenlyation. Mammalian expression plasmids are primarily used to create...create mRNA and the commonly used mammalian terminators (SV40, hGH, BGH, and rbGlob) include the sequence...commons.wikimedia.org/w/index.php?curid=17554393   Use of mammalian poly(A) signals in combination with certain viral...
  7. Illuminating Epigenetics with A FRET Based Biosensor

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    ...FRET signal back again. The Ting lab constructed 2 versions of this reporter to target the K9 (pcDNA3... 525.7570 (2015): 533-537. PubMed PMID: 26352475. 2. Bygren, Lars O., et al. "Change in paternal grandmothers...paternal or maternal copy of the gene is expressed in normal individuals (in a parent-of-origin-specific manner...because too much or too little gene product disrupts normal function. Angelman syndrome is characterized by...both cases, the other copy of the chromosome is normally imprinted and silenced, and so the syndromes are...supply influenced cardiovascular mortality of the female grandchildren." BMC genetics15.1 (2014): 12. PubMed...
  8. Plasmids 101: Degron Tags

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    ...components are 1) a SMASh-tagged protein and 2) asunaprevir (small molecule). dTAG The degradation tag (dTAG...gone through an additional round of evolution (AID-2) to reduce some of the leakiness associated with the...induced by the addition of a small molecule, and protein levels can return to normal once the drug has been ... expressed beyond the tag and small molecule. It does work in mammalian cells, mice, and even yeast, though...conditions (typically the addition of an exogenous small molecule). Degron tags can range in size from several...degrons and some of the nuances to each one.   SMASh Small molecule-assisted shutoff (SMASh) functions on the...itself when you want the protein around, leaving a minimal additional sequence (7 kDa). This system also requires...
  9. Deep Dive: Fixing and Permeabilizing for Immunofluorescence

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    ... of permeabilization and membrane integrity (Fig. 2). For example, saponin interacts with cholesterol ... at too high a concentration.      Figure 2: Examples of different permeabilization strategies... but it’s really just formaldehyde. Formalin: You may also come across formalin, which refers to a saturated...cause some confusion. Paraformaldehyde: Many IF protocols call for paraformaldehyde (PFA) as the fixative...saturated solution of formaldehyde (saturated being 37%). Formalin solutions often contain methanol as ...order to slow the polymerization of formaldehyde.  Note: Formaldehyde in solution will naturally polymerize... maintaining cellular and tissue morphology. Formaldehyde is the most commonly used cross-linker for IF...
  10. 6 Tips for Grant Writing

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    ...days before the deadline isn’t going to be much use. 2. Originality is essential for grant success The two... it’s important not to throw in the towel if 1 or 2 funding bodies say no. If possible, get feedback from...Communication Skills 5. Practice makes perfect, apply for small grants too There are actually quite a lot of different...available for researchers, so look into applying for smaller grants from within your institute or a private ...certainly found that the experience I gained in writing small travel grants to attend conferences and a seed funding...
  11. R Bodies: Membrane-Rupturing Microscopic Tools

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    ...be decorated with proteins and remain functional, 2) break membranes outside of their natural context ...PubMed PMID: 3759909. PubMed Central PMCID: PMC213417. 2. Schrallhammer, M. et al. Tracing the role of R-bodies...to find R body mutants that couldn’t extend at a normal pH and later realized that these mutations clustered...
  12. Stabilized Bacterial Promoters: Constant Gene Expression at any Copy Number

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    ...per cell increases 4-5 fold (for pUC plasmids) or ~2 fold (for p15a, R6K, and ColE2 plasmids) as cell division...biotechnology 36.4 (2018): 352. PubMed PMID: 29553576. 2. Kittleson, Joshua T., Sherine Cheung, and JChristopher...further away. While constitutive promoters drove maximal gene expression near genomic origins of replication...
  13. Pooled Library Amplifications

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    ...estimate transformation efficiency (Figure 2).    Figure 2: Example of a serial dilution to estimate... easier for a smaller, recombined plasmid to overtake full-size plasmids (or for smaller plasmids to overtake...diversity in the library is maintained. However, for smaller libraries, chemical transformations can also be...using, as well as other factors, such as your lab’s normal transformation protocol, the size of the cuvettes...direct competition with other colonies containing smaller, recombined backbones that may replicate faster...on solid media. If your plasmid library is of a smaller size with a single size insert, liquid media can...
  14. PCR Applications for Copy Number Variation Assays

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    ...mean you have one copy of the gene; an RQ value of 2 means you have two copies, and so on and so forth....quantitative PCR. Expert Rev Mol Diagn. 2005 Mar;5(2):209-19. doi: 10.1586/14737159.5.2.209. PMID: 15833050...are read after each cycle. The raw data is then normalized to a reference sequence present in the same amount...housekeeping’ gene), and quantified relative to a 'normal' sample with known copy number set as the expression... all qPCR assays, samples, standard curves, and normal controls must be run in triplicate.  The nice thing...the amount of DNA in your sample relative to your normal control - you can set up your calculations so that...RQ values used to call copy numbers should be as small as reasonably possible and set using the variance...
  15. Google Forums Round Up: First Impressions of NgAgo

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    ...engineering methods have their own pros and cons (2). The NgAgo DNA guide is inexpensive and can be 5’...Nature biotechnology (2016). PubMed PMID: 27136078. 2. Burgess, Shawn., et al. "Questions about NgAgo." ... dangerous, non-physiological temperature for mammalian cells and one which diminishes the endonuclease...PAM sequence might not always be available at an optimal distance from the target site. For example, NGG...can indeed be optimized for genome editing in mammalian cells. However, the fact that most of the researchers...rapidly degraded by the cellular machinery, leaving a small amount of active NgAgo/ssDNA complex in the cell...It is possible that, during translation, only a small fraction of nascent NgAgo protein molecules manages...
  16. Quick Guide to All Things Lentivirus

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    ..., and the desired gene (transfer vector plasmid), 2) the HIV-1 genome containing all viral genes with ...protocols that have been described (protocol 1, protocol 2, protocol 3) to find the one that will give you the...PubMed PMID: 3785217. PubMed Central PMCID: PMC367857. 2. Naldini, Luigi, et al. "In vivo gene delivery and...development of viral vectors, gene transfer into mammalian cells often relied on chemical (CaCl2, cationic...If not using proper technique, there’s always a small chance that a researcher could infect him or herself...
  17. 5 Tips for Troubleshooting Viral Transductions

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    ... digest and compare it to some reliable controls. 2. Viral stocks may be sensitive to freeze thaw cycles...24003179. PubMed Central PMCID: PMC3760253. 2. Rohwer, Forest. "Global phage diversity." Cell 113.2 (2003...An estimated 320,000 viruses can infect mammals. Even more abundant are the Earth’s estimated 1031 bacteriophages...by resuspension of the collected particles in a smaller volume. Always remove packaging cell debris before...strategy to estimate unknown viral diversity in mammals." MBio 4.5 (2013): e00598-13. PubMed PMID: 24003179...events during integration of transfected DNA into normal human cells."Nucleic acids research 18.9 (1990)...
  18. Simple CRISPR-based Epigenetic Editing: dCas9-directed DNA Demethylation

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    ...Hrit et al. 2018, Kaas et al. 2013). Figure 2: CRISPR TET-based epigenetic editing. In reality, ...approach for targeted DNA demethylation. Cell Discov 2, 16009. https://doi.org/10.1038/celldisc.2016.9.  ..., and big hair. To what was perhaps a slightly smaller population, something else was also suddenly cool...distinguishes genes of some human cancers from their normal counterparts. Nature 301, 89–92. https://doi.org...
  19. PITChing MMEJ as an Alternative Route for Gene Editing

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    ... PITCh vector via In-Fusion or SLIC cloning. Step 2: Design an insertion locus-specific gRNA The gRNA ...Find the plasmids from this publication at Addgene. 2. Dalvai, Mathieu, et al. (2015). “A Scalable Genome-Editing.... optimized the PITCh-gRNA for minimal off-target binding in mammalian genomes as assessed by CRISPR design... of alternative end-joining, requires only very small homology regions (5-25 bp) for repair, making it...similar to HDR, but it is accomplished using much smaller regions of homology, which facilitates easier cloning...
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