... from E.coli, named TadA7.10 in ABE7.10)  a catalytically impaired Cas protein (dCas or Cas nickase)  ...accidental insertions and deletions (indel) by the catalytically impaired D10A nickase from ABE7.10 can be a ...the authors created ABE8 constructs using a catalytically “dead” S. pyogenes Cas9 (dC9-ABE8.x-m/d). The...even in adult patients. In order to analyze the clinical use of ABE8 for primary human cell editing, the... reduced off-target editing frequencies. For applications that require high DNA editing specificity, for...deamination, an important attribute for therapeutic application. ABE8s demonstrate an overall improved base editing...editors with increased activity and therapeutic application. Nature Biotechnology. https://doi.org/10.1038...

... Roska’s lab at the Institute of Molecular and Clinical Ophthalmology Basel recently took a different ...Strategy #2 (ProB): Create promoters using phylogenetically conserved sequences that are upstream of the...expression. Surprisingly, <1% of synthetic promoters replicated the expression specificity of the wild-type promoters...reproducibly labeled retinal cells can be found here.  Applications for synthetic promoter AAVs Besides restricting...synthetic promoters have a few other potential applications: Using promoter combinations for targeted expression... promoters expressed in. The lab tested this application using one promoter to drive expression of Cre-mCherry... rates of labeling are ideal of gene therapy applications, promoters that produce sparse labeling would...

...progress on establishing a system for the chemical replication of RNA in the absence of evolved enzymatic...enzymatic machinery (i.e. RNA replication with no proteins). The chemistry is complex and this work requires...inheritance that might be less error prone than DNA replication. He too mentioned how “surprising” small RNAs... about using CRISPR/cas to address the growing clinical problem of antibiotic resistant bacteria. See ... and Sun Yat-sen University presented two new technical developments. Using the Tet-on system, his lab...Steitz quipped that RNA in the ribosome is in a complicated conformation “not like DNA which has a boring...

...are time-consuming, have been typically optimised for immortal/genetically-corrupt cell lines, lack scalability...alone in the cellular milieu. It is, therefore, critical to understand how proteins interact with one another... another (or with DNA) to perform diverse biochemical tasks in the cell. One of the most popular approaches... to tag the protein of interest with a small biochemical “epitope tag” such that these tagged proteins... purchased from various other suppliers with a typical turnaround time of 5-7 days. The crRNA and tracrRNA... to tag >30 genes/plate in the first shot with typical knock-in efficiencies of 10-20%. Together, our ... the non-HDR allele. For N-terminal tagging, specifically searching for error-free knock-in at both alleles...

...Because of their small size, nanobodies can be synthetically expressed off of plasmids in bacteria, and they...with a large and diverse pool of sybodies that hypothetically contain ones that bind strongly to your target...Starting with libraries and a biotinylated (for purification purposes) protein target of interest, the protocol... identifying sybodies that bind strongly and specifically to the protein of interest. The Seeger lab used...-30) of sybody candidates are sequenced for identification. All unique sybodies can be subcloned into ...2020) Generation of synthetic nanobodies against delicate proteins. Nature Protocols 15:1707–1741. https...Light to Manipulate Cell Signaling and Protein Purification RANbodies: Reporter Nanobody Fusions Plasmids...

...requires incubation at 55 °C - a dangerous, non-physiological temperature for mammalian cells and one which...been excitedly testing NgAgo's genome editing applications. So, how does NgAgo stack up thus far? A few...survey. When asked if they could detect indels (an indication of gene disruption ability) with NgAgo, one respondent...could not achieve indels with the NgAgo system indicates that it is very tricky to use and fussy to optimize...proposed in the original article, to achieve greater efficacy, researchers will have to carry out repeated transfections...results out there include: Google Groups - I've specifically set up a forum where NgAgo users can share protocols... S. Lander, and Feng Zhang. "Development and applications of CRISPR-Cas9 for genome engineering." Cell...

...McDonald. "A practical guide to evaluating colocalization in biological microscopy." American Journal of... proteins of interest and examine them using biochemical methods thus removing the context of the cell...way it is important to choose the appropriate quantification methods and tools. It is usually not enough...microscopy is limited by the wavelength of light and, practically speaking, for common lab microscopes the detection...Quantitative analysis of dynamic association in live biological fluorescent samples." PloS one 9.4 (2014): e94245...

... (green or blue) within a population of timer indicates the age of the pool: the higher the ratio, the...fluorescence falls within the linear window for the theoretical timer shown in Figure 1B, you can use the equation...-FT-N1 pTRE-Slow-FT pBAD/HisB-Slow-FT -  Applications of fluorescent protein timers FP timers have...protein, or cellular structure and a particular biological event (trafficking to a subcellular location,...of neural fiber bundle formation." Journal of Biological Chemistry 276.32 (2001): 29621-29624. PubMed ...to red report on cellular trafficking." Nature chemical biology 5.2 (2009): 118-126. PubMed PMID: 19136976... insulin-secreting beta-like cells." Nature communications 7 (2016). PubMed PMID: 27063289. PubMed Central...

... most commonly used are GECIs - genetically encoded calcium indicators (e.g. GCaMP). Others include: pHluorin...capacity. Many of these FPs are fluorescent when ectopically expressed in most organisms. Furthermore, fusing...fluorescence. Therefore, FPs are used to study many biological questions. The two most common uses are: 1) to...For LSS proteins, the Stokes shift is ≥ 100nm. Specifically, these proteins are excited by UV light or blue...were prone to oligomerize. This may affect the biological function of the FP-fusion protein. Therefore,.... al. (2010) “Fluorescent proteins and their applications in imaging living cells and tissues”  Physiol...

...excited to announce we now offer an export license application for rabies materials! Some of the materials in...international requestors. We also offer export license applications for VSV and SARS-COV-1 plasmids. Export control...will need to provide a brief description of the applicable research project when purchasing export-controlled... applied to cover processing costs. Once the application is approved, Addgene then ships the material ...offered VSV and SARS-COV-1 plasmids under this application process since May 2020. We find this process ...whenever possible, as we cannot expedite the application process.  More information about export control...