... that uses fluorescently labeled Cas9 to label multiple genomic loci in live cells. While other systems...multicolor labeling would enable the visualization of multiple loci. Such a system could then be used to examine...synthetic scaffold that can be used to recruit multiple protein molecules (and also available from Addgene...could also have major applications to cancer, for example, the visualization of chromosomal translocations...

...Collecting that much tissue takes a week of multiple people performing dissections. With CUT&RUN, we have...and cell lines. Although ChIP-seq is a relatively simple and robust protocol it does have limitations. The...CAPTURE Additional Resources on Addgene.org Find simple lab protocols Browse CRISPR plasmids for isolating...

...well as developing yeast as source for dietary supplements! Development of a bacterial expression system...plants.     Transgenic yeast as source of dietary supplements Dr. Mrinalini Nair, University of Baroda, India...alternative source of essential omega-3 fatty acids complementing vegetable oil, a somewhat poor natural source...them. By using yeast integration vectors (see for example this yeast cloning kit) she intends to make yeast...

...process you would like to monitor, there may be multiple steps you could examine, i.e. for an enzymatic...given ion - we’ll use calcium biosensors as an example. Many calcium biosensors include GFP fused to calmodulin...reticulum (ER), vacuole or cellular membranes. Examples include blue flame mitochondria-targeted calcium...available, and the conditions of your experiment. For example, does the fluorescent tag need to be pH stable ...

...give you the reverse sequence, complement sequence, or reverse and complement sequence.  Filter DNA tool:... tools help with that.  DNA sequence reverse complement tool: Enter a DNA sequence and the program will...DAD): This tool designs overhang sets for high complexity Golden Gate Reactions Cloning ligation reaction...

...Suitability for in vivo experiments, due to their simple structure    sdAbs in research These advantages...becoming valuable tools for diverse applications. For example, RANbodies can be used as an alternative to conventional...manipulate cellular functions, like in this early example we highlighted back in 2015, where sdAbs were used...around using sdAbs in diagnostic applications. For example, fluorescently and radioactively labeled sdAbs ...

...see how people describe their actual work. To do this, try looking at LinkedIn profiles of people who have...experience.  Use LinkedIn to see what types of jobs people with master's have Job descriptions often fail ...the LinkedIn search filters to limit results to people that work in industries you’re interested in (i.e...

...awareness of FOP. Please consider running with us or making a donation via the link below. Please continue to...visit us and watch fellow Run4Rare team members complete the Boston Marathon, he wanted to run and play...on October 23rd. This is a great event at which people dress up like superheroes or other iconic characters...

...of being a successful manager. Great bosses tell people where they stand clearly and routinely. They are...feedback when it is merited. Surprisingly, some people find this hard so get used to it by making it a...for a feedback session by writing down specific examples and notes so you can get back to the issues if...problems. While giving corrective feedback isn’t pleasant, it’s far worse for employees if you don’t care...

...is left to do with the characterization and implementation of this system. Monday evening's talks switched...foremost, from the custom software used to design complex genetic circuitry to the downstream output. Tuesday...kept busy for the whole poster session, talking to people from every conceivable background at the meeting...new CRISPR/Cas users) and updated Synbio page to people who were curious about these newest features of...

...probably good news for our environment that more people are paying attention to living a “green” life and...pile up by the sink after lunch as fewer and fewer people use plastic bags.  We also try to recycle our discards..., but I never let that stop me. I only had to replenish my glass milk supplies every 10 years or so. Actually...less stop buffer and miniscule amounts of plasmid sample.  How satisfying?  With practice, I got good at...

...listen for hints at the same concerns from multiple people. Listen to what they say, and what they don...grad school vs. postdoc Choosing an advisor to complete a successful PhD is different from choosing a ...confidentiality to ensure more honest answers. Ask people who work in the labs next door for their opinions...

...genome engineering tool offers flexibility, multiplexibility, and ease of use. In order for this technology...protospacer adjacent motif (PAM). Once the CRISPR/Cas complex binds to the foreign DNA, a cut is made to destroy...the tool’s specificity, orthogonality, and multiplexibility in various species and describe an assortment...from experts in the field, and enabling access to multiple plasmids used for various platform applications...

... sequencing and RNA sequencing of complex heterogeneous RNA samples. Furthermore, such high processivity...ever like to write about a recent plasmid deposit please sign up here. Listen to the Hot Plasmids Podcast...technologies based on polymerase processivity. Please find the MarathonRT plasmid here! Zhao C, et al...

...Collaborations may be less “complicated” - for example, easy to initiate a collaboration without having...matrix - can be frustrating for solo flyers Other people will routinely scrutinize your data—science can...issues can create interpersonal issues (one bad apple can change entire culture) Specific requirements...Can move between the two (helps to know the right people) -> How can you make a decision about something...

...method kills the transgene free individual. For example, with CRISPR-Cas9 gene editing, transformation ...GUS staining or are not suited to studying large samples. RUBY Red But recently, something new has been ...belongs to a different chemical family than the red/purple plant pigments found in all other plants, anthocyanins...Other innovative uses of RUBY are possible, for example as positive selection of transformed plants to ...

... Biosensor GENIE Project New viral prep with multiple serotypes pAAV-hSyn-GRAB_OTmut AAV1 Biosensor...Biosensor Robert Campbell New viral prep with multiple serotypes AAV-hSyn-Ribo-jGCaMP8s AAV1 Biosensor... AAVrg Control Karl Deisseroth New serotype (multiple) rAAV-CAG::FLEX-rev:: hM4D-2a-GFP AAV1 Chemogenetics... Multiple Authors...

...interact during various behaviors. Either tissue samples were harvested after the behavior, or the animals...adaptable tool that could continuously monitor multiple systems. Ideally this tool would be wireless, ...develop these fibers, the labs looked at previous implementation of optogenetic tools, calcium sensors, and ...optical stimulation of its associated fibers across multiple channels and can also record tissue temperature...

...Viruses are simple: a genome packaged in a protein shell (Taylor, 2014). They’re so simple that we can... even decide if they’re alive or not. Yet these simple, small particles have quite the outsized impact... been genetically modified to either limit or completely eliminate their replicative ability. The remaining...

...The triple T’s of biology are transformations, transfections, and transductions! In this blog we will...expressed. Once the design and construction steps are complete, it’s time to make virus! The constructed viral...the biological outcome you want to achieve? For example, do you want the nucleic acid cargo to be integrated... to achieve different biological outcomes. For example, some viruses integrate into the host genome (lentivirus...

...modular, selection-based gene tagging approach Multiple features of C. elegans have made CRISPR/Cas9 so...HDR mentioned above, an accessible germline, and multiple selectable markers. Bob Goldstein’s lab previously...The data are clear - the SEC-based system is both simple to use and highly efficient for C. elegans genome...elegans protein. The his-72 locus is shown as an example. Used with permission from the Genetics Society...

..., in turn, the big picture of the project. For example, how can future regulations impact your work? What...word “extracurricular” may be taboo in the lab, multiple research studies highlight the importance of taking...potential careers to gather more information Follow people in academia, industry, and government on social...sounds interesting, TAKE ACTION! Taking action: Examples of extracurricular activities The table below...

...cellular level. The reason for this is cost and complexity – anyone who works in the lab knows how expensive... engaging for students. In our BioBits kit for example, we’ve designed FD-CF experiments that result in...educational activities using the BioBits platform, please get in touch with us! We’ll work with you to get...Addgene.org Browse our protocol videos to learn simple lab techniques Check out our molecular biology ...

...practical and hands-on. Students in these courses get plenty of theory in their lectures, but it can be difficult... format I decided on was to have two teams of 4 people face each other on a single chosen clinical disease...modify BUILD YOUR PLASMID or generate games like it, please freely share your projects with us! This exchange...want to receive our modifiable powerpoint file, please feel free to contact us at: [email protected]...

...you can fully investigate and solve a problem or complete a project, overcoming inevitable failures and ...Technologist/Designer     Fieldwork and traveling Many people are attracted to science because they enjoy spending...shows that you are passionate and capable. For example if the job involves traveling, showing that you...organised and experienced traveler by citing specific examples, may help you get your foot in the door. Match...

...accelerating research into this virus. One such example is cell lines. The American Type Culture Collection...global endeavor. Case study 2: Plasmids Another example of reagents that have broad applications but have...dozens of times in 2 weeks. This is a striking example of the contributions that scientists are able to...popularity of the hACE2 transgenic mouse strain, for example, is sustained by the availability of this strain...

...ul) samples of EGFP-expressing AAV packaged in various serotypes. This collection is now complete! pAAV-CAG-GFP.... Likewise, a lot of scientists have requested samples of AAV for testing purposes. To address this, we... kit with all the serotypes, since these 20 ul samples are distributed individually. See our Serotype ...

...JA. Precise transcript targeting by CRISPR-Csm complexes. Nat Biotechnol. 2023 Jan 23. doi: 10.1038/s41587...only having one binding site per protein. Having multiple antibodies allows you to use them in parallel ...recombinant monoclonal antibodies for enhanced multiplex immunolabeling of brain. Elife. 2019 Jan 22;8:...  Multiple Authors...

...major histocompatibility complexes (MHC class I and II).  There are multiple algorithms, both open-access... antibody drugs are popular therapeutics for a plethora of disease conditions, from cancer to autoimmune...approach The in silico approach cannot replicate the complexity of antigen processing events that take place ...

...emotional time!” George Church tries to recruit people who are nice as top criterion and offers advice...productive in a handsoff environment. “Treating people fairly does not mean equally.” George Church was...comments that it is also surprising “ how easily people talk themselves out of great projects as ‘impossible...building up your lab and selectively picking the first couple of hires to build a strong, collaborative team....

...for plasmid replication in mammalian cells, for example, and a toxin that kills bacteria may not have any... is possible that you may not want the plasmid depleted, but still want to use transient transfection ...HeLa, NIH3T3, CHO, COS-1, 293HEK, Jurkat T cells Complexes with DNA; causes strand scissions 100-400 ug/mL...your cells divide. Gentamicin is often used as a supplement in mammalian cell culture to suppress bacterial...

...was not at all the case. The conference featured people who work in sociology, behavior, and artificial...versa. Even within the biological sciences, there’s plenty of interaction between neuroscience and other fields...microscopes, and dissection tools, but there were also people ready to discuss their VR helmets, gait tracking...field Not to toot our own horn too much, but many people who came to the Addgene booth (there were over ...

...2004. We’ve already distributed over 500 viral samples to scientists all over the world. Now that the ...successfully launched, I would like to thank some of the people and organizations who helped us reach this milestone...donations. That seed money was used to build a completely self-supporting service so that Addgene is not..., tracked production and distribution of viral samples. The Office Team made sure we could ship internationally...

...stable host cells (the simplest example would be a transformed mesophilic, for example E. coli, carrying a...potential collaborators and interested audiences. Please visit our team wiki or contact us at igemiiserb...

...molecular biology and scientific research. There are multiple ways foreign DNA can be introduced into cells ...plasmid can give them different phenotypes. For example, this plasmid encodes a green fluorescent protein...eukaryotic cells via a virus or viral vector. One example are bacteriophages that attach to bacterial membranes...References Baltrus, David A., and Karen Guillemin. "Multiple phases of competence occur during the Helicobacter...

...conditions in order to maximize the mutation rate. For example the Phage Assisted Continuous Evolution (PACE) ...diversity introduced by lambda red recombineering and Multiplex Automated Genome Engineering (MAGE) will always...interest. A gRNA is used to direct the PolI3M-nCas9 complex to a DNA site of interest, which nCas9 nicks and...progeny of a single starting cell. Additionally, multiplexing of guides allows EvolvR to target more than ...

...primer known as a megaprimer. Piecing together multiple PCR products using sequence homology is also common... Cloning, and SLIC can all be used to assemble multiple fragments, but these methods can be expensive ...could make the E. coli do the work by using the principles of recombineering. In recombineering, bacteria...other methods, such “BAC shaving” would require multiple steps. One potential pitfall of REPLACR-mutagenesis...

...this complex holds a pair of DNA ends together, forming a paired end complex The paired end complex then... factors can also influence NHEJ activity. For example, repressing RecA and overexpressing the NHEJ machinery...

...promoter region, which is predicted to be required to decouple gene expression and plasmid number. In a suite...fluctuations. When cells are dividing rapidly, multiple rounds of replication are initiated before the...greatly impacting the level of gene expression. For example, the synthesis of deoxychromoviridans requires ...products is needed, such as within a multi-protein complex, stabilized promoters can eliminate variability...

...ever like to write about a recent plasmid deposit please sign up here. No time to read? Listen to our Hot...aberrantly activate the proteasome. Clift et al. tested multiple methods for TRIM-Away delivery. For small cell...rapid and reversible recruitment of chromatin complexes to any genomic locus in any cell type. By varying...FIRE-Cas9, experiments can be designed to target multiple loci at once. In addition, the approach allows...

...minutes of inactivating the CASANOVA complex, fluorescent CRISPR complexes were observed binding telomeric ...ever like to write about a recent plasmid deposit please sign up here.          Listen to this episode ...2015) or ratiometric Bimolecular Fluorescence Complementation (rBiFC, Grefen and Blatt, 2012). The 2in1 Plasmid...

...accessible, anyone could present the material. For example, Addgene’s contributions by Tyler Ford included... and matched to serve a specific audience. For example, I joined this effort later in the year and in ...diverse and international team assembled the right people at the right time. Be inclusive and seek solutions...like to join or contribute to this collaboration, please check out the official website, follow the group...

...service, and more For the first time in the last couple of years, we don’t have to worry about having enough...community is loving it. Almost 10,000 viral vector samples were distributed in 2018 and we are poised for ...unsolicited praise: “Addgene, for one, is a sterling example of collaboration and innovation between legal and...Our long reaching reputation helps us hire great people from farther away via connections and even via ...

...plasmids between scientists. The service would allow people to add a gene to a repository others could request... had 1,000 depositing scientists. Ensuring that people knew how to access Addgene, and that we were making...which continue to grow today. “I generally refer people to Addgene for the basics of molecular biology ...reagents, and we realized we could use the same principles to provide recombinant antibodies, another plasmid-based...

... when scientists talk about their precious RNA samples. RNA is inherently less stable than most macromolecules...RNA. To protect the RNA from degradation, several simple methods have been employed which modify the sugar...suggest coupling several of them together, for example phosphorothioates with 2’-O-methyl modifications...RNA to control the entire editing system with a simple LED light. Lighting up your gRNA with fluorescent...

... channels and pumps in developmental biology.  Examples of some ion-channel and ion-pump encoding genes...genes HCN1 KCNJ2 KCNJ13 GABRG1 KCNIP2 KCNQ1 Examples of some genes encoding gap junction proteins cx43...also modulating bioelectric states in vivo. For example, the Levin lab from Tufts University has used Arch...sensitive fluorescent voltage indicator with near-complete desensitization in continuous light. ArcLight ...

...on the symposium as a whole and for registration please go to http://futureofresearch.org/boston/. The ... about how the system operates and functions” “Complete lack of information on number of postdocs” “Postdocs...comments are well correlated with published data. For example, in 2010, less than 15% of postdocs had achieved...beginning to address these issues independently; for example, Harvard’s Therapeutics Graduate Program has a ...

... into functional multi-subunit macromolecular complexes, but a serendipitous discovery led them to investigate...different Arg-mutant enzymes could be rescued by supplementation with guanidine hydrochloride (GdnHCl). Rescue...previous approval by the FDA, GdnHCl could, in principle, provide an effective treatment for rare diseases...concentrations (≤3 mM) of GdnHCl can functionally rescue multiple Arg-mutant proteins in living baker’s yeast cells...

...EnvA receptor (TVA) in user-defined cells. For example, a researcher can deliver Cre-dependent TVA to ...optogenetic or chemogenetic receptors) enables complex control of specifically defined neuronal populations...how newly generated neurons integrate into the complex pre-existing circuit of the brain. Another study...circuitry, and continue to contribute answers to the complex questions of how the brain operates. References...

...myself. I like being with the people and for the people, feeding the people, and making them get outside...enjoy the field I am in, the atmosphere, and the people.” My absolute favorite response to this question...