...Virus Genome Interactions Important for Vector Production and Transduction. Human Gene Therapy 31:499–511...and Rep40) Rep proteins. Alternative splicing generates the different C-terminal regions of the Rep proteins...transfer plasmid, which contain regions of dsDNA. To generate a 3’ hydroxyl group needed for DNA replication...in the ITRs, uses its endonuclease activity to generates a ssDNA substrate which is then nicked by its ...splicing and differing translational start sites generate the three VP proteins. The three VP proteins assemble...Adeno-Associated Virus (AAV) as a Vector for Gene Therapy. BioDrugs 31:317–334 . https://doi.org/10.1007/...Improving the Efficacy of AAV Vectors. Molecular Therapy - Methods & Clinical Development 12:248–265 . https...

...genome engineering - such as certain forms of gene therapy - avoiding the use of viruses is also important...lab screened many SB tranposase variants for hyperactivity in mammalian cells. Their efforts resulted in...approximately 100-fold more efficient than the first-generation transposase (as determined by transposition assays...construct. This new tool kit allows the rapid generation of stable transgenic cell lines which can robustly... CD19-directed chimeric antigen receptor (CAR) therapy. SB is a valuable tool and an alternative to CRISPRs... Optimized Sleeping Beauty transposons rapidly generate stable transgenic cell lines. Biotechnol J. 2015...Mates L, et al. Molecular evolution of a novel hyperactive Sleeping Beauty transposase enables robust stable...

...through the proliferation of specific, and not all, Langerhans cells, to create foci of proliferating cells....Brainbow The Lichtman group developed the original iteration of this technology in 2007. Used to label neurons... is expressed. Cre-mediated deletion of one or several of these genes permits the expression of genes ...occurs. Following the development of Brainbow 1.0, several more refined versions have been developed, which...tracing in zebrafish with Zebrabow Before the generation of Zebrabow (Pan et al., 2011), Brainbow was ...Tissue-specific Cre recombination in this mouse generates cells that randomly express either nuclear GFP...become a powerful tool for understanding cell proliferation. Inducible XFP expression in the Ubow mouse ...

...guide RNAs may tolerate a mismatch in the binding sequence, we recommend changing several bases as close... to as ‘indels’. While this is convenient for generating gene knockouts, NHEJ repair does not allow introduction...introduction of specific sequence changes. To generate a specific sequence change the cell must undergo...other groups have focused on strategies to improve overall HDR rates, we have shown recently that desired ...desired genome-editing events by HDR can be generated more efficiently by (1) improving editing accuracy by ...systematically characterize this phenomenon and found a general relationship between the distance of a mutation...probabilities our data revealed, and found that for generating a homozygous HDR event with optimal efficiency...

...edited. For many experiments, the end goal is to generate a clonal cell line, so it’s important to verify... for a needle in a haystack before going on to generate and screen individual clones. If a clonal line...al., NAR). This technique uses the trace file generated in Sanger sequencing and a simple algorithm to...with your sgRNA sequence. The TIDE software will generate a graph representing all insertions and deletions... 4 clones. Did you use a dual guide system to generate your knockout? Then you’re in luck! You don’t ... to visualize an edit. Most dual guide systems generate a deletion large enough that it can be visualized...pair edit with a prime editor. What next? When generating a knock-in mutation, the best screening method...

...which chews back the 5’ ends of the fragment, generating long overhangs which allows the single stranded...stranded regions with homology to anneal a polymerase, to fill in the gaps a DNA ligase, to seal the nicks...enzymes is that they can all work at the same temperature, so the entire reaction takes an hour or less...advantage is that this process makes it easy to generate wild type and mutant constructs at the same time...an overlap of 15-40 bp, with a primer melting temperature greater than 48℃. Both Snapgene and NEB have ...instead of using the exonuclease feature of T4 DNA polymerase, which must be controlled by the presence or ...methods that have been described--CPEC (circular polymerase extension cloning) and SLiCE (Seamless Ligation...

...important consideration is whether the affinity of an experimental protein-ligand interaction approaches...non-specific binding). Historically, glutathione-S-transferase (GST) fusion proteins have been immobilized on... target protein with the vitamin biotin. The interaction between biotin and the egg-white protein avidin...affinity (i.e. sub-nanomolar) protein-ligand interactions. Avidin and its derivatives have four biotin-binding...involves amine-reactive biotin derivatives that generate covalent bonds with positively-charged amino acids...solution was to exploit the best of both worlds. We generated a plasmid that produces biotin ligase and a protein...erythropoietin receptor and subsequently evaluated interactions with mutant and wild-type erythropoietin molecules...

...we believe it is best to further reduce onsite operations for the coming week. In an abundance of caution...Addgene charges enough for our services to run our operations. We don’t raise money except occasionally for...to launch a new service or product. Our daily operations for all ongoing processes are entirely self-supporting...they grasped the crucial role Addgene plays in accelerating research. Yet this was still not enough to ensure...funding organizations to request support for our operations until our normal work resumed. Addgene distributes...type of biomedical science research. Our work accelerates and enhances productive collaborations as well...distribution of plasmids for COVID-19 research is accelerating essential research in laboratories all around...

...Over the past several months, we’ve seen the emergence of several SARS-CoV-2 variants coincide with increased...the world” says Balazs. “By combining those with [sera] we got from people that have been given the vaccine...relative ability to neutralize.”   Figure 2: Sera used in the study came from individuals who received... two doses of the Pfizer or Modern vaccine. This sera was used in the high-throughput SARS-CoV-2 pseudovirus.... In their assay, the pseudoviruses express a luciferase reporter which is used as a readout for infection...Figure 3: Neutralization outcomes of convalescent sera with various pseudotyped virus. Image from Garcia-Beltran...quick to deposit their plasmids at Addgene to accelerate ongoing COVID-19 research. “I hope that by sharing...

...The contents of this post are intended to convey general information only and not to provide legal advice...you think about your career (and your future in general), keep in mind that you may have years to work ...for recent Ph.D. graduates). Therefore, it is imperative to plan for and develop your career into a series...Order which sets broad policy intentions directing federal agencies to propose reforms to the H-1B visa system...collaboration management, grant administration, clinical operations, and so on. Candidates should look broadly at...and your continued work is necessary to create therapeutics to cure the cancer. Authorship:  Publishing ...Interest of the EB-1A. Furthermore, the ability to leverage the prestige of the employer makes EB-1Bs a very...