...efficiency of multiplex gene editing. Moreover, existing multiplex strategies have been facing difficulties... fused base editors (nCas9-BE), as Cas9 is mechanistically more favored for efficient base editing. They...human cysteine tRNA stood out from their initial testing. It outperformed other tRNA candidates (either ...multiplex genome engineering of crops or allow for sophisticated CRISPR screenings that would perturb the unprecedented... only minimal genetic payloads. Different from existing multiplex array architectures, including previous...

... also be used to engineer new pathways with sophisticated gene expression controls. Most cell types do...two-component systems (TCSs). TCSs consist of a histidine kinase that phosphorylates a response regulator...domain from cyanobacteria to a common E. coli histidine kinase. This hybrid construct senses light, and...light; these two systems can be coexpressed for sophisticated control of gene expression. An E. coli light-sensitive...light-sensitive two-component system (TCS). A hybrid histidine kinase/photosensor senses various light inputs...

...when seeking your dream lab. The principal investigator's point of view If you are looking to join a ...course, the specific research field should be interesting; however, this should (in most cases) not be ...be the driving factor. The following questions can serve as a guide for the quest of a perfect "fit". ...Scientists". This post also includes a useful list of questions for a candidate to ask lab heads, lab members ...human disease. For more details and many more suggestions on planning for future success by finding and...

...reach out to the program coordinator with your question(s). This precautionary step increases the likelihood...communicates your interest in a specific program, and institution. Consequently it’s best to draft a cover letter...vast majority of programs tend to ask similar questions, thus devoting some time to writing a solid personal...interview. Formulate and practice a set of common questions with a peer or mentor. Several days prior to the...entry-level position following graduation. While these suggestions increase your competitive edge, they alone, are...

...Biology. Can you tell me where to start?” is a question more and more students are asking. Teachers are...of educational programs all focused on current questions and experiments in the field of synthetic biology...from her own MIT curriculum. Drawing on current questions being asked in MIT’s research labs, BioBuilder... into modules that students and teachers can investigate together. Teachers decide which BioBuilder lessons...ideas ranged from a biological breathalyzer to pesticide breakdown, production of fire-resistant housing...

...Since there are many thousands of partitions, statistical modelling can be used to determine the actual...mismatch nuclease assays (2, 6, 7). Ability to distinguish between mono-allelic and bi-allelic mutations...digital PCR assays is that they are easily able to distinguish single-cell derived samples with mono-allelic...instructions on how to design such assays, in addition to testing their performance (2). Remember that although digital... digital PCR is a great screening tool, it still requires target DNA sequencing to confirm the exact nature...

...or +1 (617)-ADDGENE (233-4363) with additional questions. Bacterial Strain Addgene can distribute your...Return to Article" button to return to the table listing all of the plasmids for this deposit. You can then...deposit, associate the plasmids with a Principal Investigator (PI), provide us with your shipping address ...hereby certify that I am affiliated with the institution ___. If this is no longer true, please modify...material under the terms deemed necessary by my institution or other third party rights holders. Once this...January 2016. It was updated by Angela Holmes and Christina Mork in February 2024.  Additional Inside Addgene...

...researcher at Lunenfeld-Tanenbaum Research Institute, posting about Addgene’s 20th anniversary on Twitter...SirHamhands, responding to a request for “any suggestion for CRISPR online course for beginners” on r/...alike. We transformed this process by aligning institutions with standardized agreements and electronic ...!” — Julien Béthune, Professor at HAW Hamburg, posting about his Blue Flame Award on Bluesky. In 2016,...

...cytosine and adenine base editors that can be reconstituted in vivo to form a protein identical in sequence... plasmids and co-transfected into cells for reconstitution. Image from Levy et al., 2020 with permission...when paired with appropriate fluorescent optics. Existing red fluorescent voltage indicators, however, fall...neural circuit manipulations when combined with existing green-fluorescent GEVIs and blue-light-sensitive...The kit functions in the model cyanobacterium Synechocystis sp. PCC6803, and the fast growing strain Synchococcus...

... the fragments size of your digestion on an agarose gel. If the digestion does not provide the fragments... to your cloning strategy: Restriction enzyme digestion Be mindful of the restriction enzymes you chose...like XbaI, ClaI, etc. If you are doing multiple digestion be sure that the buffers and temperature are compatible...It's also best to use 1-2 ug of the vector for digestion. PCR Design primers that overlap at least 24 bp...

...antibody region that binds to the antigen) from existing antibodies. Each cleaved antibody will produce...(Muyldermans S., 2013). They have the unique distinction of being the only affinity reagent on this list...  To antibody or not to antibody?  That is the question… or is it? This post isn’t meant to convert anyone...trendy as that may sound. Validated antibodies are still an excellent option for many applications, and frankly...References Grumezescu, A. M. (2018). Drug targeting and stimuli sensitive drug delivery systems. Elsevier Science...

...available to counteract the toxin. If, however, an investigator were to successfully clone an insert into the... (re-ligated empty vectors, for instance) would still express ccdB and therefore would die. This procedure...insertion will completely replace ccdB with the investigator’s insert of interest. Hence correct clones are... behind the “swapping out” of ccdB with the investigator’s gene of interest.   CcdB resistant E. ...further. Feel free to offer your comments and suggestions, and even to submit a guest blog entry of your...

...controlled by deliberate and compulsory actions instigated by the central nervous system signaling to peripheral...metabolic syndrome. Coregulators Coregulators constitute a group of some 400 molecules that have been ...regulated manner. The fact that coregulators belong to distinct complexes may explain how more than 300 different...collectively address many important and fundamental questions in nuclear receptor and coregulator research. ...the NURSA website will feature direct links to listings of available plasmids for our curated NRs and ...

...how close is close enough for the desired edit to still occur if the cut site is sub-optimal? As with everything... cut site is just 10 bps away from the site in question (Kwart et al, 2017). To add insult to injury, ...same PAM preference of NG was isolated by the Kleinstiver Lab (read more on this lab's other developments...Nonetheless, in an AT-rich region, these Cas9s may still leave you choosing between efficiency and location...location.   Nearly PAM-less editing   The Kleinstiver lab sought to address the PAM problem through structure-guided...

...penicillin resistance plasmid can grow in spite of a hostile Penicillium fungus trying to kill them off. While...resistance to toxic substances like heavy metals or pesticides, enabling their hosts to survive where others... This ability to endure a harsh environment is distinct from the benefit of resistance plasmids, which...shifted over the years, and new classifications are still being created as whole-genome sequencing becomes...Plasmids Isolated from Lactic Acid Bacteria:Their Characteristics and Potential Applications in Biotechnology...

...certain cell types.  Botond Roska’s lab at the Institute of Molecular and Clinical Ophthalmology Basel ...finding retinal cell-type specific promoters. Most existing promoters express in a broad number of cell types...targeting the same cell class in all three species are still greater than random chance. The chances of a promoter...human retinas but not in mice was low (R=0.12), suggesting that mice could be used to eliminate promoters...Morikawa, R.K., Rompani, S.B., Hantz, P., Szikra, T., Esposti, F., Cowan, C.S., Bharioke, A., Patino-Alvarez,...

...ball-and-chain that they resemble, actually impede your investigation into the regulation of your favorite gene. But...these studies report correlations without clearly testing whether a certain methylation state causes a condition...to create rare DNA bases that are recognized by distinct proteins and result in unclear transcriptional...achieved using lentiviral backbones expressing blasticidin or puromycin resistance genes.  It’s that simple...Feinberg A, Vogelstein B (1983) Hypomethylation distinguishes genes of some human cancers from their normal...

...a risk that they may underestimate viral titer as these assays cannot distinguish between a cell that ... based off of p24 quantification tend to be overestimates since they can include free p24 and defective...assay, titering via measurement of viral RNA can overestimate the amount of infectious virus in a prep as ...integration events, reducing the possibility of underestimating the viral titer. The main drawback of this ...

...engineered precursor of the systemic capsid in question (or both). Therefore, a high-efficiency systemic...These are only starting recommendations; you will still need to experimentally determine the best titer ...your experiment, it’s time to start selecting and testing systemic capsids! Figure 2 contains a decision ...reduced efficiency, and is not neutralized by pre-existing antibodies against AAV9-based vectors (and vice...AAV-PHP.S, with a bias toward segments of the small intestine (Chen et al., 2022). Systemic capsid for alveolar...Cre-dependent selection yields systemic AAVs for targeting distinct brain cell types. Nature Methods, 17(5), 541–550...

...in 5’ four base overhangs after cutting. The destination vector receives the assembly after similar Type...re-digested and "eligible” for ligation into the destination construct. Reactions are often cycled between...acquisition of antibiotic selectable markers by the destination plasmid. Antibiotic selection allowed for high...very occasional uncut or cut/re-ligated pGGA destination plasmids. A final validation of the 5-, 12- or...is dependent on the presence of every module, destination construct and functioning Type IIS restriction...