...cloning) is a DNA cloning method that does not use restriction enzymes or ligase, and requires no post-PCR procedures...depicted as a green cloud) functions both as a restriction endonuclease and as a ligase by cleaving and ...Design standard primers (no need to add unique restriction sites on the ends) and amplify your sequence ... confirm the presence of your insert by PCR, restriction digest, or sequencing. Pro tips Do not add 5...

...This post was contributed by guest bloggers Lauren Celano of Propel Careers and Rachel Casseus, Esq. ...practice in that field. H-1B work-authorization is strictly limited to employment by the sponsoring employer...of the work of those in your field. Original contributions to the field that have had a major impact. For...of original scientific or scholarly research contributions in the field Evidence of authorship of scholarly...Addgene.org Careers at Addgene Global Plasmid Distribution ...

...of proteins for degradation using AID Article contributed by Alyssa Cecchetelli Auxin Inducible Degradation... of CRISPRCas9 by engineering SpCas9 Article contributed by Shreya Vedantam CRISPR/Cas9 is widely used...channelrhodopsins for the optogenetic toolbox Article contributed by Susanna Bachle Anion-conducting channelrhodopsins...31346176. Using LADL to loop with light Article contributed by Eric Perkins Mammalian genomic DNA is not ...

...This post was contributed by Manon Eckhardt and Melanie Brewer from the QBI Coronavirus Research Group... these proteins are FDA-approved or in clinical trials or preclinical studies already. We are following...all viral genes are flanked by EcoRI and BamHI restriction sites - so cloning projects should be fun and...partnering with Addgene, who will be taking over the distribution to expand the network of scientists working ...

...This post was contributed by Deborah Sweet, Vice President of Editorial at Cell Press. Almost everyone...Communication is the issue Much of the problem can be attributed to communication, and in particular communication...(accession numbers, reagent details, author contributions, etc.) before their paper can be published. ...across life and physical science, and editorial contribution to the strategic development of Cell Press. ...

...This post was contributed by guest blogger Lydia Morrison from New England Biolabs. What is DNA assembly...standardized DNA constructs, but its use of Type IIS restriction enzymes results in scarless assembly. Finally...allowing fragments to be taken directly from restriction enzyme digests to DNA assembly. What makes it...containing regions of homology) cut from plasmids via restriction enzyme digestion, and used directly in DNA assembly...

... exploded so quickly, citing the first clinical trial with iPSCs with his collaborator Takahashi in 2014...perspective: Dr. Yamanaka talked about his clinical trial patients with advanced macular degeneration; Laurenz...presented work that is leading to promising clinical trials for ALS.   I have an uncle with Parkinson's, a ...first stem cell line that could be used in clinical trials. The quality control process alone takes nearly...

...also use electroporation, the application of an electrical charge to cells, to increase cell membrane permeability... integrated phage remains dormant until it is triggered to enter the lytic cycle.   During both of these...they observed genetic recombination between two nutritional deficient E. coli strains that resulted in a ...cells. Electroporation on the other hand uses electrical pulses to create transient pores in the cell ...

...floats freely, depicted in the figure below. Once triggered by blue light, the CRY2 and CIB1 proteins heterodimerize...specificity as full-length Cas9 (flCas9). When triggered by blue light (470nm), paCas9-1 induced indel ...response to far-red light (730 nm). This nucleotide triggers the translocation of the protein BldD to the nucleus...constructs. This proof-of-concept model opens intriguing possibilities for optogenetic manipulation of...Gene editing Manna et al., 2019   NPPOC-caged trimethoprim (TMP) Cas9 fused to destabilizing domains (Cas9...

...resources. There are three types of cognitive load: intrinsic cognitive load is the effort associated with a...even the most enthusiastic of learners.  What contributes to cognitive load?  Everything! Here are some...specialized knowledge and context I have spent years striving to obtain?” If the answer is yes, you will need... or to gloss over some aspects when needed. The trick is to communicate your key points well enough that...

... on size is ultracentrifugation. Sedimentation equilibrium analytical ultracentrifugation yields an affinity...This post was contributed by guest bloggers Joachim Goedhart and Marieke Mastop from the Section of Molecular...on the dimerization tendency in living cells. Strikingly, the OSER assay reveals that tagRFP does not ...

...palindromic repeats separated by an 8-bp long asymmetric core spacer sequence. The asymmetry in the core...described above, a genetic modification can be restricted based on experimental constraints. This has been...site have been created, which contain a unique asymmetric spacer "NNNTANNN", where "N" indicates which ...system in your research? Do you have any tips or tricks, or clever cre-dependent techniques?  Let us know...

.... The main challenge with using AdV is that it triggers a strong immune response after in vivo administration...personalized medicine (2017).   1st Gen AdVs are stripped of regulatory genes E1 and E3. Without these genes...plasmid stock should be analyzed by endonuclease restriction digest or fully sequenced to confirm its integrity...chloride (CsCl) density gradients combined with ultracentrifugation to separate rAdV from other cellular debris...

...into the liquor of the cerebral ventricles (ICV, intracerebroventricular) to minimize tissue damage; however...Experimenter Interaction should be minimal and strictly controlled; optional handling animals Experiment...depending on the choice of channel protein, they can trigger neuronal excitation or inhibition upon light (or...

...data (Begley & Ellis, 2012; Baker, 2015). This contributes to an enormous amount of wasted money and time...Nguyen K, van der List D, Murray KD, Engebrecht J, Trimmer JS (2019) A toolbox of IgG subclass-switched recombinant...https://doi.org/10.1038/520295b Bradbury ARM, Trinklein ND, Thie H, Wilkinson IC, Tandon AK, Anderson ..., Bestagno M, Burrone O, Maynard J, Ferrara F, Trimmer JS, Görnemann J, Glanville J, Wolf P, Frenzel A...

...that the five traditional rules didn’t need to be strictly followed in order to generate high-fidelity reactions...overhang set by uploading the set with the Type IIS restriction enzyme and thermocycler protocol. It will also...a circular or linear assembly. If you aren’t constricted to a specific number of fragments, you can also...correctly assembled phage genomes, it required a very stringent selection process and benefited from a longer ...

...in any format. Distribution status for your plasmids: Hold for Publication or Distribute pending QC. The...Gibson cloning, ligation-independent cloning, restriction enzyme, TOPO cloning, unknown, or other. ...or licenses that could prohibit Addgene from distributing the plasmid, select “yes” from the drop-down...

...This post was contributed by guest bloggers Keith Pardee and Alexander A. Green. Zika background First...successful sensors to draw on as our assays grew more stringent, and it avoided any days of lost productivity ...when we needed additional expertise. Perhaps most striking is that there were seven institutions involved...and open approach by all involved definitely contributed to our ability to complete the project on such...

...antibodies against proteins embedded in membranes is tricky because they have fewer hydrophilic surfaces, require... randomized with a defined set of different trinucleotides (corresponding to different amino acids) in...exchange (FX) cloning technique which uses Type IIS restriction sites (Geertsma, 2013). Once the phagemids are...after a few washes, binding is detected using a colorimetric reaction. After the ELISA step, the narrowed-down...

...This guest post was contributed by Sana Khan Khilji, a PhD student at the Max Planck Institute of Colloids...that UV light only kills microorganisms that it strikes directly. UV light exposure is also harmful for...way to prevent contamination. This sounds rather trivial, but often solves most problems before they occur...PubMed Central PMCID: PMC4047077. 2. Pagliara, Patrizia, et al. "Ostreopsis cf. ovata induces cytoskeletal...