...time spent at the desk, but it is worth it. This requires a focus on deliverables. It does not mean a manager... and colleagues Sometimes this means you have to fire a good performer if they are enough of a jerk to... of most people, not just scientists) Are self-directed, but need precise leadership and support from ...Scientists. Subscribe to the Addgene Blog to follow the entire series as it is published. Good introductory reference...

...the recognition domains—including the CTD and the entire REC lobe—do not structurally resemble any other...weak, random binding to precision gene editing requires three major steps: RNA loading, DNA sequence recognition...The RNA guide sequence is extended as a single unpaired strand in the center of the protein. The repeat...1, nestled between the REC and CTD domains, is required for Cas9 nuclease function. Stem loops 2 and 3...But how to test DNA bases that are already snugly paired to each other? At this point, the protein can switch...

...and varying methods of replication. In fact, an entire classification scheme, known as the Baltimore replication...follow central dogma (gDNA to RNA to proteins) or require an intermediary DNA replication step between DNA...virus. RNA genomes can be transcribed into mRNA, directly translated into proteins, or, in the case of retroviruses...reproduce when experimentally necessary, genes for the required proteins are packaged in separate plasmids and...

...tagmentation preparation approaches described previously require a preliminary double stranding step. The double...single-stranded and both TA-based ligation and tagmentation require a double-stranded template.  Using viral genome...of serotype confirmation, this method does not require any additional reagents or lab work (Pacouret, ...polynucleotide chains. J Virol 9(2): 394-396. Lecomte E, Tournaire B, Cogné B, Dupont J-B, Lindenbaum P, Martin-Fontaine...

...this awesome experiment you want to do, but it requires some AAV. You’ve never worked with AAV before,...plasmids have a promoter, but this element isn’t required when the transfer plasmid harbors non-coding sequences...’s talk about some of the elements that aren’t required parts of an AAV transfer plasmid, but are often... control transgene expression. Lox sites are directional 34 bp sequences that can flank the transgene ...

... my curiosity did not ultimately translate to a desire to do the research itself. Several years of exclusively...more creative faculties into motion, such that a desire to once again write for fun eventually saw an English...remarking to a classmate that, if this was one day required of me for work, I would have a high level of career...neurological health risk. Having played the sport my entire life, I was curious as to what the research had...

...wide variety of applications. But some techniques require you to select against the presence of a transgene... Antibiotic selection would only then kill the desired plants! PCR or fluorescent markers are currently...plants, but screening these can be challenging and requires specialized equipment. Reporters for studying ...from these plants will be green, containing the desired mutation but lacking both RUBY and the construct...

...researcher at the Wyss Institute for Biologically Inspired Engineering and member of the Addgene Advisory...production frustrating - a researcher can complete an entire lentiviral production protocol only to find that...widely requested and incredibly useful tools, they require extensive background work from the requesting scientist...Church lab at the Wyss Institute for Biologically Inspired Engineering and the Zhang and Doench/Root labs...

...choosing a FP, but much of the primary data one might require when making such a decision remains spread across...to us for advice on choosing a protein, we often direct them there as a starting point.  However, there...previous versions.  Currently, all submissions also require a DOI referencing primary literature, and every...information on a particular FP; this can be done directly using the search bar on the homepage (and at the...

... favorite books - both scientific and not - to inspire you to read a little more this October. Science...science of race."     - Joanne Kamens, Executive Director  I Contain Multitudes by Ed Yong "Microbes are...another secret."     - Marcy Patrick, Associate Director of Quality Control Jonathan Strange & Mr. Norrell...Club (ABC) founder, Eric Perkins, is Addgene’s Director of Science Product Management and explains the...

...frequency. Although highly useful, this technique requires extensive sample processing to avoid disrupting...appear red. To enable this color switching, TRICK requires the following components: a GFP protein that ...field of view for constant tracking. You can then directly monitor multiple rounds of translation from this...caveats. In both cases, creating a reporter mRNA requires inserting multiple hairpin sequences into the ...

...population. The benefit of beads is that they do not require cells from your experiment, and they are non-specific...., CD4, CD8b, CD11b, CD19), are less likely to require an FMO. Markers that define more specific cell ...often used in the later gates, are more likely to require FMOs. Isotypes Isotype controls are negative controls... populations, but others do not. Some journals require isotype controls, while others don’t. However, ...

... nucleus through nuclear pores and thus do not require breakdown of the nuclear envelope during cell division...gamma-retrovirus are so closely related, their required components are very similar. While plasmids containing...specific cell type doesn’t always lend itself to direct application of virus. However, the cargo capacity...infect dividing and non-dividing cells and doesn’t require genomic integration to express its cargo, in fact...

... AAV variant that efficiently infects neurons, directs high-level transgene expression, and has retrograde...any genetic tool delivered by the AAV.   AAVrg requires high titers for efficient retrograde transport...dye-based neuronal tracing reagents, AAV particles require receptor-mediated entry into the cell. This means...inject a recombinase-dependent chemogenetic vector directly into the region where the cell bodies of the projection...

...used. All of these expression control mechanisms require a specific modification of the viral vector for...of interest, for example. Some customization is required, but many viral constructs have already been built...applications. These libraries include sgRNAs targeting the entire genome, overexpression of all human transcription...delivery of the actual virus to which immunity is desired. Instead, components of the virus are now delivered...

...blessed to have an incredibly supportive Board of Directors at Addgene. Our Board helps keep our focus on ...best practices and encouraging us to explore new directions. I would like to thank them for their time, dedication...can afford to do and when. Addgene’s Legal Team acquired permission from relevant plasmid depositors and...proud to be adding viral vector services to our repertoire. Additional Resources on the Addgene Blog ...

...ancient Chinese game Go). In addition, there was an entire story-telling session organized by Story Collider...in neurobiology (and really in all of biology) require large data sets. Whether these data sets include...or sequences of microbial DNA they all seem to require some mathematical know-how for analysis. If you...from Addgene. While super useful, these vectors require a bit of expertise and can be difficult for individual...

...students in real-world lab activities? I’m trying to inspire them and also keep up with all the new biotechnology...being asked in MIT’s research labs, BioBuilder inspires students and teachers to apply engineering principles...breathalyzer to pesticide breakdown, production of fire-resistant housing insulation, and cleaning up salt...BioBuilder's open-access curriculum available on Amazon Inspire.    Many thanks to our guest blogger, Mary Tamer...

...membrane so DNA can be introduced directly into the cell. Microinjection requires the use of a fine needle to... bacterium’s genome through homologous or site directed recombination. Transduction is a common tool used...donor bacterium to a recipient bacterium through direct contact. The donor bacterium contains a DNA sequence...

...(FP) require external light activation, you can’t use fluorescence to monitor processes directly affected...luminescent imaging: A luminescent substrate is required for imaging, and it is important to verify that...technology. Multicolor imaging using Nano-lanterns requires a linear unmixing algorithm to separate the various...

...is not enough on its own to help you find your desired plasmid. In many cases, you will need a screening...you will need to sort through them to find the desired clone. Do your selection, then screen Why add a...what about identifying clones that contain your desired insert? This is where a  screen comes in. Let’s...process. This way, we can confirm the sequence of the entire plasmid. Please note that NGS verification is more...

... paper for publication in a scientific journal requires several considerations. First, it is important...; the reviewer’s name and affiliations, and the entire referee report are permanently attached to the .... If the reviewers suggest that the manuscript requires revisions, the authors can publish a new version...with the journal’s house style and check that all required information is provided. The clearer you write...

... “make the ask” once you have found someone you admire and want to learn from. Check out Joanne's Reddit...learn. Mentors can be senior to you or peers you admire.  If you see someone who gives a great talk, ask...path you are interested in following? Someone you admire for their insight and way of expressing it in meetings...locally. Your university probably has an alumni directory. Alumni who enter their information in these searchable...

...programmed to bind to essentially any DNA sequence desired. This technology allows you to create a pair of...knock out a gene. One can also exploit homology-directed repair by flooding the cell with a mutant template...to build a DNA-binding domain to recognize any desired 15 base pair sequence in the genome in rapid fashion...to create zinc finger nucleases that will bind desired sequences. TALENs are much more reliable. If you...

... and allow you to design additional hypothesis-directed experiments. As you might have guessed from their...For some libraries, plasmid DNA can be delivered directly to the cells of interest. With others, notably...

...secondary antibody you should use when performing indirect immunoassays. The Fab regions, on the other hand...hand, are the “arms” of the Y and comprise the entire light chain and a variable and constant region of...to detect a Fab fragment, it either needs to be directly conjugated to a reporter or you need a light-chain...specific conjugated secondary antibody. These requirements limit the flexibility of and demand for Fab ...

...different mechanism thought to be associated with bidirectional transcription. In an uninfected cell, dsRNA ...interspersed nuclear elements can generate dsRNA by bidirectional transcription or as inverted repeats. These ...genomes which can produce dsRNA as a result of bidirectional transcription of their circularized DNA. These...dsDNA molecule, it does so at the expense of the unpaired DNA strand. A ssDNA molecule exists within the...

...contributed by guest blogger, Gary McDowell, executive director of Future of Research. On December 1st 2016, many...salaries - in some cases reversing these decisions entirely. At Future of Research, we had been collecting...are going ahead with raises anyway, including the entire faculty of the biochemistry department at the University...McDowell! Gary S. McDowell is currently the Executive Director of Future of Research. He is interested in advocating...

...In a recent law review article we received some entirely unsolicited praise: “Addgene, for one, is a sterling... 3.5 Addgenies strong. The Addgene Board of Directors is mature and poised to support our growth. Thanks...outgoing Chair) and Dave Root (incoming Chair) and our entire supportive Board, we have a robust committee process...referrals. Our long reaching reputation helps us hire great people from farther away via connections and...

...is the output. The input, plasmid copy number, directly influences gene expression. By itself, increasing... the promoter region, which is predicted to be required to decouple gene expression and plasmid number... example, the synthesis of deoxychromoviridans requires three genes whose proteins are needed sequentially...ideally be optimized to promote generation of the desired product while avoiding the accumulation of toxic...