.... What aspect of GeoCas9 are you most excited about? Thermostability? Longevity in human plasma? Something...cocaine esterase engineered to have increased thermostability also had a longer lifetime in human blood. ...lower than optimal temperatures. GeoCas9’s thermostability could also be used with in vitro molecular ...CRISPRing! References Harrington, Lucas, et al. "A thermostable Cas9 with increased lifetime in human plasma... thermostable cas9...

...post. There’s so many great tips for getting the most from your conference experience so we unfortunately...them all. What are your top tips for getting the most out of your scientific conference? Resources on...

...reproducibility. Read on for some tips for getting the most out of your lentiviral transduction experiments....point to consider is how the virus is being titered. Most titering approaches transduce into highly permissive...

... other Addgene news Find the most recent plasmid blog posts Find the most recent CRISPR blog posts Resources...

.... Our most popular kit of the year was the Golden Gate TALEN Kit, which continues to be our most requested...distributed through Addgene over our first 10 years. 10 most distributed plasmid technologies pSV-PGC1 - Ok,...Fire's set of vectors for C. elegans research was our most requested item of 2005. shRNA - In 2006, we saw ...two plasmids from Didier Trono's lab are our two most popular plasmids of the past 10 years. Induced Pluripotent... Yamanaka's lab and James Thomson's lab were the most frequently requested. Ubiquitin - One particular...interest in from 2008 to 2009 was ubiquitin. The most requested ubiquitin plasmids were pRK5-HA-Ubiquitin-WT...

...wanted to celebrate our depositing PI’s who have the most highly requested plasmids. Currently, there are ...

...yeast chromosome. Listen to learn more about Dr. Ellis, synthetic biology, synthetic chromosomes, and ...

... the Quake Lab. Identifying IgnaviCas9, a hyperthermostable Cas9 This year it was wonderful to learn that...Ignavibacteriae phylum genome. IgnaviCas9 is a hyperthermostable Cas9 CRISPR nuclease that is active up to ...optimal sgRNA spacer length of 22-25 nts, and is thermostable between 31-100°C. They published their finding... in a bioRxiv article and in PNAS. Two other thermostable Cas9 enzymes have previously been characterized...ThermoCas9, IgnaviCas9 is a type II-C Cas9. IgnaviCas9 is most similar to a type II-C Cas9 from Parvibaculum lavamentivorans...What can you do with IgnaviCas9? Why would a thermostable Cas9 be useful? It can enable applications where...steps in the protocol. IgnaviCas9’s incredible thermostability makes the enzyme particularly useful. Additional...

...recommend it except in the most dire of circumstances.) Multiplex assays Mo’ proteins, mo’ problems may not be...region, where an antibody binds to its target. While most secondary antibodies recognize broad isotype classes...across applications. Western blots, for instance, almost always use the indirect approach, while flow cytometry...assay and protein of interest is often one of the most challenging components of any immunoimaging assay...incorrect. With the exception of flow cytometry, most immunoimaging assays do not allow for absolute quantitative... and occasionally ask themselves and others that most perplexing of questions: “who moved my goat anti-rabbit...

...created and designed for the Hall of Human Life at the MOS. (If you want a fun refresher on the Central Dogma...scientists, part of the Addgene family Perhaps the most heartfelt portion of the entire night was when Melina...

...work in your lab. Find the FLINC biosensors here. Mo et al. Nat Methods. 2017. PubMed PMID: 28288122. ...

...Susanna Harris, Heather Hendrickson, Jonathan Jacobs, Mo Kaze, Christina Kellogg, Heather Martinez, Pat Schloss...don’t have the networks or experience to get the most of meeting.  I experienced this at the ASM General...and helping societies help their members get the most out of their meetings. Amanda Gunn ran randomized...

...Bieniasz PD, Jäger U, Porter CD, Friedman T, McClure MO, Weiss RA (1997) Sensitization of rhabdo-, lenti-...so commonly used is that it is good at infecting most cells, but not all. For example, LDLR expression...envelope proteins for pseudotyping While VSV-G is the most commonly used envelope protein for pseudotyping,...

...genome. They have four chromosomes: the X (and Y), second, third, and fourth chromosomes. Also, the fourth ...third chromosomes are the ones you’ll work with. If YFG is somehow on the Y or fourth chromosomes, you ...geneticists use balancer chromosomes to prevent genetic recombination. Balancer chromosomes essentially are messed...the start of the day and throughout daytime. They mostly stop emerging at night. An incubator with a light...fourth and Y chromosomes are miniscule, so researchers generally do not manipulate them. Essentially, the X... messed up chromosomes with multiple inversions. They are unable to participate in genetic recombination... viable if they are homozygous for a balancer chromosome. Thus, you can stably maintain a heterozygote...

...tropicalis, the two most commonly used species of frog. (Liu et al., 2016). Some of the most popular tools ...emerging model organisms!) Escherichia coli One of the most recognizable and utilized model organisms is Escherichia... organism for over 60 years with a status as the most studied prokaryotic organism. With its fast growth...low cost, it’s understandable why E. coli is the most popular bacterial organism in microbiology and biochemistry...developed to thrive in laboratory settings. One of its most attractive features is its stability and easily ...Youn, 2016). For these reasons rats were once the most used mammalian model in medical research with an...has its perks, it is far from perfect. One of the most limiting attributes of rats is their difficult to...

...vasculature (adult marmosets), biased toward astrocytes (adult rhesus macaque) Rats, marmosets, green monkeys... including ENS (adult marmosets, infant rhesus macaques) Mice, rats, marmosets, rhesus macaques Unknown... screened in marmosets. AAV.CAP-Mac effectively transduces vasculature in adult marmosets (Chuapoco et...transduction levels in marmoset brain, with both neurons and astrocytes targeted. While marmoset liver transduction...capsids for your research? While systemic capsids mostly function like your average AAVs — and have many...accommodate larger, modular genetic payloads. Since most systemic vectors are primarily derived from AAV9...

...two different pairs of chromosomes using gRNAs specific to sequences on chromosomes 9 and 13. They next ... fluorescent in situ hybridization (FISH) and chromosome capture suffer from low resolution and can’t ...flexibility of CRISPR has improved our ability to target almost every genomic locus, and to do so in vivo. Catalytically...puncta observed. By labeling two genes of a given chromosome simultaneously, they could also monitor the spatial...distances from 2-75 Mbp. Labeling multiple, intrachromosomal loci Building on the work of Chen et al., ...turned their attention to mapping pairs of intrachromosomal loci. The technique successfully resolved ... this work represents the first mapping of intrachromosomal loci, a major benchmark in characterizing ...

...Trends: Sequence based input is by far the most common with most tools also requiring that the user specify...CRISPR software embodies solutions that facilitate most experimental applications in CRISPR engineering....can invest our time into learning and extract the most value? This is only one of many decisions faced ...are two major areas where software can help. The foremost handles the design of sgRNA and represents the... suitability. Functions of this category are the most common between tools. Examples: “single-target design...design”, and “single-method design”. Trends: The most common utility functions are batch design or multiplex...detect off-targets. The few offline tools available mostly work on any computer but generally require working...

...ligase is a NAD+-dependent DNA ligase from a thermostable bacterium that can survive high temperatures...advantage of the high fidelity of Taq and other thermostable DNA ligases. These applications detect specific...amplified fragment) and treated with a high fidelity thermostable DNA ligase. If the target sequence is present...helix shape (22). Large purine:purine mismatches and most smaller pyrimidine:pyrimidine mismatches are typically...several DNA ligases bound to substrate (23, 24). Thermostable DNA ligases, including Taq DNA ligase, are naturally...detectably ligate many T:G, T:T, and A:C mismatches. Thermostable DNA ligases are active at elevated temperatures...below). Conclusion It is important to note that thermostable, high fidelity, nick-selective DNA ligases like...

...two different pairs of chromosomes using gRNAs specific to sequences on chromosomes 9 and 13. They next ...example, the visualization of chromosomal translocations or chromosomal shattering (chromothripsis). As...catalytically dead Cas9 (dCas9) has many applications, the most well-known being transcriptional activation/repression...could then be used to examine both inter- and intrachromosomal dynamics, and how they change in response ...turned their attention to mapping pairs of intrachromosomal loci. The technique successfully resolved ... this work represents the first mapping of intrachromosomal loci, a major benchmark in characterizing ...next! References Multicolor CRISPR labeling of chromosomal loci in human cells. Ma H, Naseri A, Reyes-Gutierrez...