...unable to reach all regions of the CNS. PHP.B, a neurotropic variant of AAV9, is better than AAV9 at gaining...receptor used by PHP.B viruses. Creating PHP.B, a neurotropic AAV9 variant PHP.B’s specificity for C57BL/6J...CNS transduction and transduces the majority of astrocytes and neurons across multiple CNS regions. PHP....Ly6a seemed the most likely candidate due to its strong linkage to the PHP.B phenotype and its expression...mice strain specific Ly6a SNPs on PHP.B binding in vitro. They found that when LY6A contained C57BL/6J SNPs...PMID: 30819613. Hordeaux, Juliette, et al. "The neurotropic properties of AAV-PHP. B are limited to C57BL...

...most as cancer cells are amenable to culture in vitro. Virologists too have benefited from the ability... ‘Master’ stock, which can be stored in liquid nitrogen long term, preferably some of these vials will...will be stored in a separate backup liquid nitrogen store to prevent loss from equipment malfunctions. One...also monitor the stock levels you have in liquid nitrogen for your Working and Master stocks of each cell... laboratories to carry our independent quality control for a range of testing, from animal doping to forensics...Thermofisher Guidelines for Maintaining Cultured Cells Invitrogen Cell Culture Basics Guide Abcam Cell Culture ...

...are available to confer greater transcriptional control, assist with proper protein folding, and deal with...Contains mutated lac permease whch allows for linear control of expression F- ompT lon hsdSB (rB- mB-) gal ...polymerase (RNAP) from the bacterial genome under control of the lac repressor, which is inducible by the...cases, some strains carry an additional measure of control such as the pLys plasmid, which suppresses basal...difference being that the latter provides tighter control of basal expression. What if I don't see protein...sources, so fear not—there are a few simple troubleshooting measures that can help get you back on track...

...Emerging Investigators Podcast breakdown Intro Chapter 1 - Introduction to Mark Springel, Olivia Ho-Shing, ... In this episode of the Addgene Podcast, we introduce you to the Journal of Emerging Investigators, an...developing one-on-one mentorship programs. Chapter 6 and Outro - Interview with high school student researchers...basic plasmid biology Use or CRISPR 101 eBook to introduce students to this amazing genome enegineering technology...

...Yeast Prototrophy Kit from Markus Ralser's Lab -  Use this kit to complement unused auxotrophies in Saccharomyces...to a quick (~5 min) Hot Plasmids podcast that introduces you to 4 new plasmid technologies from one of... Plasmid 2: Magneto 2.0 form Ali Guler's Lab - Control neuronal activity with a magnetic field. Listen...strains.  Listen to Our Segment on The Yeast Prototrophy Kit Special thanks to our sound engineer/producer...

...coverage, mutational efficiency, control over where mutations are introduced, and scalability (see Supplementary...favoring of certain types of mutations, difficulty controlling the rate of mutagenesis, and low throughput. ...mutagenesis PCR that uses mutagenic primers to introduce all desired codon changes (step 1 in the image... high-fidelity Phusion polymerase are used to introduce point mutations into the newly synthesized DNA...Error-Prone PCR”. Methods in Molecular Biology In Vitro Mutagenesis Protocols, pg. 103-109. PubMed PMID:...

...Shine-Delgarno sequences used in this so-called bicistronic design (BCD). The 49 vectors in this kit can ...of varying strengths centered within GFP/RFP bicistronic reporters characterized in Cambray et al., 2013...terminators can be used to precisely tune your polycistronic E. coli expression constructs. Some applications... Living Bacteria Using an Orthogonal Ribosome-Controlled Fluorescent Reporter."ACS chemical biology 11.1...vector that enables protein expression in any in vitro translation system. The gene set was carefully chosen...been used to produced recombinant proteins for in-vitro assays: Koch, Daniel, et al. "A pull-down procedure...

..., such as the introduction of heterozygous endogenous epitope tags, or the introduction of dominant mutations... examples from their lab include using AAV to introduce epitope tags into the endogenous alleles of the...2008). They have also used AAV gene editing to introduce naturally occurring cancer-causing mutations into...However, while CRISPR is remarkably effective for introducing non-sequence specific frameshift mutations through...efficiency of AAV-based gene editing via the introduction of a double strand break by using a targeted...

... the strength of the signal in comparison to a control sample. Southern blotting applications range from...causes sickle cell anemia. The mutation happens to destroy a sequence recognized by the restriction enzyme...gels and next-generation sequencing techniques Introduced in 1977, Sanger sequencing, also known as chain-terminating...referred to as sequence DNA as it synthesized by electronically recording fluorescence activity as fluorescently...differences in ion flow caused by the different electrochemical properties of nucleotides (8). Compared to ...sequences among DNA fragments separated by gel electrophoresis." Journal of molecular biology98.3 (1975): ...

...A few years ago, researchers introduced what’s been called a major breakthrough in cloning technology...plant model systems. “The cloning is all done in vitro; it’s no different in mouse, human or plants,” explained...of an advanced molecular plant science lab.” Introducing GreenGate As Lohmann and his colleagues describe...describe in the PLoS ONE paper that introduced the kit in December, GreenGate includes the six most frequently... available through Addgene,” Lohmann said. “We strongly believe the free sharing of material is absolutely...

...and did their best to claim the costume contest trophy. We’ve had teams dressed up as plasmids, Blugene...unilateralis is a fungus that infects ants and controls their behavior. These ants become “zombies” and...their jaws into a leaf, and dies as its muscles atrophy. After the ant’s death, the fungal fruiting bodies...display imitating a moving caterpillar. The worm controls the snail’s behavior, forcing it out of hiding...blood out. Once the blood is depleted, the tongue atrophies and falls off. The louse remains becoming the ...

... in a pooled cell population, as mutations with strong phenotypic effects can mask weaker ones. Thus, ... the need for a faster and higher-throughput in vitro approach. Using data from The Cancer Genome Atlas...them had not yet been functionally validated. In vitro assay allows first-time functional annotation of...collection to screen individual perturbations in vitro using Ba/F3 and MCF10A cellular models. Ba/F3 and...effect on cell viability compared to a wild-type control. Approximately ~22% of the mutations, however, ...

...afflictions. Elia Di Schiavi - Spinal muscular atrophy Dr. Elia Di Schiavi is a Research Scientist at ...Naples, Italy. His group works on spinal muscular atrophy, a disease in which disruption of the smn1 gene...the smn1 gene is known to cause spinal muscular atrophy, the molecular mechanisms behind the disease are...restore motor neuron function in spinal muscular atrophy patients. Noushin Nabavi - Peritoneal Mesothelioma...of incidence and prevalence of spinal muscular atrophy (SMA)." European Journal of Paediatric Neurology...

... The results are then visualized using gel electrophoresis. Surveyor™ and T7E1 have been widely adopted...research setting, but these events could have disastrous health effects in the clinic. NGS-based methods...including Digenome-seq (Kim et al. 2015) for in vitro analysis, IDLV for in vivo detection (Gabriel et...potentially toxic) Digenome Seq 0.1% Cell-free (in vitro) Works across all cell types Must be verified with... background BLESS Not reported Cell-based (in vitro) Can be used on tissue from whole animal models,...most investigators use single cell clones for in vitro CRISPR experiments. The likelihood that a single...

.... Layers of control have also been added to the CRISPR technology to allow time-controlled edits to occur...target can be identified if it isn’t known. If a strong candidate emerges, downstream optimization is performed...generation of knockout cell lines which has made this control step extremely accessible. For example, let’s say...  Susanna Stroik...

...traps can be used as high-throughput screens to introduce reporters throughout an organism’s genome. These...Enhancer traps do not need to be integrated within an intron or exon of a gene, and thus they do not disrupt...splice acceptors. They require insertion into an intron of a gene, where they can be spliced into the gene...Additional resources on Addgene.org Vector database Intro to the Lab Bench Plasmids 101 e-book Additional... Susanna Stroik...

...of input material, cells or tissue, to produce a strong enough signal over background noise. This is not...access to transcriptional regulatory elements is controlled during development using the Drosophila wing ...the target protein. This targeted digestion is controlled by the addition of calcium, which MNase requires...only for a short period of time, thus precisely controlling the amount of cutting and mitigating noise generated...protocol, the most significant being the usage of a strong detergent to permeabilize cells rather than to ...

...ways budding young scholars across Europe get introduced to the cultures and opportunities available to... societies to promote their upcoming events and intro sessions, and give out useful items and information...fairs are also where many future scientists are introduced to the biology and chemistry societies that will... to attend. We thought we’d take some time to introduce you to the idea of the Freshers Fair and some ...Scientific societies like those mentioned above provide strong foundations upon which young scientists can build...

...are similar to transfections, processes which introduce foreign nucleic acid into cells by any means, ...safety considerations, they aren't dangerous for controlled lab usage. Using viral transductions can offer...selected to perform the transduction with: either retrovirus, lentivirus, adenovirus, or AAV. Then, your desired...integrate into the host genome (lentivirus and retrovirus) while others are non-integrating (adenovirus... Susanna Stroik...

...recombination of bacterial plasmid DNA: electron microscopic analysis of in vitro intramolecular recombination. ...One of the most frequent problems that I help troubleshoot involves viral vectors and plasmids with repeating...testing your prep can save you the headache of troubleshooting failed experiments. Pick the right strain While...backbone.   Ready, set, grow! Our 5 Tips for Troubleshooting Viral Transductions will help with the next...