...addition to certain staff scientists and those in technical roles. Therefore, institutions are responsible...suggested that the salary level should now be automatically adjusted every 3 years. In May 2016, Secretary...centrally administered, which leads to many complications, including the inability of some institutions...paid. Institutional salary policies are one indicatator of how salaries may changes due to the FLSA. ...from $43,692 to $48,216, an increase of 10.4%. The Icahn School of Medicine at Mount Sinai in New York is...of the postdoctoral workforce. Based on our communications with and research on these institutions, we...to the new level:  We are still waiting for an indication of what (most of) Harvard, Johns Hopkins, (most...

...on DNA, or are enzymatically active. For this reason, recombinant proteins are typically expressed in E...expression vector cannot have p15A origin of replication F- ompT lon hsdSB (rB- mB-) gal dcm (DE3) pLysS...expression vector cannot have p15A origin of replication F- ompT lon hsdSB (rB- mB-) gal dcm (DE3) pLysE...Expression vector cannot have p15A origin of replication F- ompT hsdSB(rB- mB-) gal dcm (DE3) pRARE2 (...Expression vector cannot have p15A origin of replication fhuA2 [lon] ompT gal (λ DE3) [dcm] ∆hsdS/ pLemo...Expression vector cannot have p15A origin of replication F-, Φ80ΔlacM15, thi, lac-, mtl-, recA+, KmR Expression...medium. Induction timing is important, as you typically want to make sure your cells have first reached...

..., Nature Communications, 2019. PubMed PMID: 31451697. Genetically encoded voltage indicators for use ...Article contributed by Brook Pyhtila Genetically encoded voltage indicators (GEVIs) change fluorescence in ...lentivirus and have been difficult to manipulate genetically. To solve this issue, Sarah Teichmann and colleagues... many aspects of ApoB-LP biology, including quantification of abundance, size, and localization in vivo...Pubmed PMID: 23068583 Thierer et al, Nature Communications 2019. Pubmed PMID: 31366908 Adapting sigma54...real time when neurons or muscle cells transmit electrical information. Such voltage imaging can be performed...measurements such as patch clamp recordings. However, technical difficulties such as small fluorescence changes...

...for post-translational modification of recombinant protein products, specifically glycosylation. The native...scene. But what about the other yeasts – in the biomedical world, are there even other yeasts? In this blog...2017). Beyond human relevance, yeast have many applications in food science and biotech due to its fermentation...The most commonly used yeast by biotech and pharmaceutical companies is K. phaffii (formerly known as ...the system. Since then, multiple biotech and pharmaceutical companies (along with some research laboratories... bread. The following table lists some of the biological commonalities and differences between K. phaffii... ancient yeast strains and metazoan cells.   Applications for K. phaffii There’s a number of different...

...than the classical purification protocol involving iodixanol gradients. The current purification protocols...particles accumulate inside the producer cells and purification protocols often describe a process that begins...lysis to release AAV particles with subsequent purification steps. It’s also possible for AAV vectors to...from Massachusetts General Hospital and Harvard Medical School took a closer look at the AAV vectors that... This is particularly useful for therapeutic applications in vivo where endogenous anti-AAV antibodies...antibodies often neutralize the therapeutic efficacy of AAV vectors. How are exosome AAVs produced? Purifying ...exo-AAVs seem to target cell types with a similar specifically as conventionally produced AAV particles. But...

...taraxacifolia (Willd), an annual herb of Western Tropical Africa typically known as wild lettuce. Aside from their...diabetics nephropathy, one of the major pathophysiological complications of diabetes mellitus. Recombinant ...researchers in resource-limited settings. More specifically, the Addgene-Seeding Labs plasmid grant provide...formation. The quest for plant compounds to treat complications of diabetes Gloria Otito Izu, PhD Candidate...flow. Novel treatment options, as well as non-pharmacological solutions to modify the course of this condition...scanning microscope will allow Tsakanova to image cortical neurons and study changes in neural activity after... employing synthetic biology resources and microfabrication technologies to generate a self renewable ...

...To our surprise, we measured identical dissociation rates for catalytically inactive dCas9 molecules. We...altering its function). HDR is crucial for certain applications, for example, correcting the allele that causes...these editing reactions is low. Understanding the biological cause of this repair bias is a fascinating (and...(Richardson et al 2016) revealed some of the biophysical parameters that can influence the HDR/NHEJ decision...key scientific principle: that understanding a biological process in detail often suggests new strategies... dissociation rates and discovered that the catalytically active form of the nuclease dissociates very...Our in vitro results also demonstrated that catalytically inactive dCas9 produced a “bubble” structure...

...to actually hold all those seemingly abstract biological reactions that I learned about from textbooks...physics class to learn about electronics or mix chemicals together in chemistry to see what happens, but... expensive tiny amounts of reagents are, how complicated (and again, expensive) the different machines...proteins outside of a cell - cell free extracts Typically, if you want to make a protein – say green fluorescent...proteins without the need to handle a living, replicating organism. Furthermore, cell free systems can ...they can see, enzymes that can convert certain chemicals into fragrances that they can smell, or enzymes...central dogma while also giving them practice with practical lab skills like developing a hypothesis, precisions...

...together can inform decisions about in vivo application and clinical testing. This would pave the way for improved...helps researchers understand a drug’s safety and efficacy in vivo.  ADAs and Adverse Immune Reactions Anti-drug...drug and increase drug clearance, lowering its efficacy and increasing the risk of an adverse immune reaction...silico approach The in silico approach cannot replicate the complexity of antigen processing events that... evaluation of ‌therapeutic immunogenicity is typically performed using traditional 2D assays with immune...drug product while ensuring the safety and the efficacy.  Better together Combining in silico and in vitro...This system would somewhat act like a 2-step verification. All approaches have their own limitations. ...

... has been critical to the Broad’s success over the last 11 years and was certainly critical to drawing...background in history and specifically the history of RNAi left him a bit skeptical of the technology at the...cancer biology in Ed Harlow’s group at Harvard Medical School and then applied to work at the Broad Institute...towards the Broad. In order to ask interesting biological questions, you need a tremendous amount of technology... in true collaboration with scientists asking biological questions. Doench loves that he still gets to...terms of determining what genes are involved in biological process X, you can do with CRISPR technology....with Addgene - he says he’s “known of Addgene basically since the beginning and [has] been using Addgene...

...ITRs serve as a type of origin of replication to initiate replication of the dsDNA virus. Viral packaging... endogenous E1 gene, making it replication deficient. This is typically supplied by the production cell...plasmid while the other plasmids with packaging or replication components are called helper plasmids. Below,...viral experiments is to deliver a genetic cargo, typically an expression construct, to cells/organisms. This... production, they can be helpful for certain applications and aid in ease of use. Our lentiviral guide...yield dsDNA. Unlike LTRs, ITRs are perfectly symmetrical to allow for this pairing to occur. Like LTRs...AAV are rep and cap. Rep is required for viral replication and also aids in infrequent genome integration...

...addition of a helper virus or plasmid allows it to replicate episomally, essentially remaining an extrachromosomal...extrachromosomal part of the host genome that can replicate and divide along with the endogenous DNA. AAV also...human gene therapy and is currently being used in clinical settings. AAV is also a cool tool in the laboratory...cell type doesn’t always lend itself to direct application of virus. However, the cargo capacity of AAV ...gamma-retro viruses. This is large enough for many applications, but in some cases multiple AAVs (co-infection...that we’ve reviewed the common viruses used in biological research, you might have an idea which ones could...and AAV. If you’re wondering more about what applications these viruses are good for, we have plenty of...

...promising therapeutic applications; it can enhance therapeutic delivery across broader physical areas of the brain...driven by neurons. By studying how neurons are physically connected, we can begin to understand how the...Therefore, by simply looking at how neurons are physically connected we can begin to understand how the ...region (anterograde tracer, Fig. 1B). Tracers are typically delivered to a particular location and they either...neuronal signaling, or manipulating any other genetically controlled function. By using AAVrg in Cre transgenic...Cre-dependent AAV into Broca’s area of a mouse specifically expressing Cre in the hippocampus (Fig. 3). ... pigs (work you can find in AAV Data Hub!) The cortical pathways to the IC were well transduced by AAVrg...

...with biological protocols and revolutionary physical approaches. However, with each new technical achievement...aim to make microscopy as a tool accessible to practically everyone.     Fig. 1: Open source hardware...convinced that most microscopy techniques can be replicated with commercially available consumer-grade components...modify and optimize the code, find and fix security-critical bugs, and contribute to constant updates of the... OpenFlexure microscope or open-source modular optical toolbox openUC2 are trying to overcome this hurdle... tools has allowed the community to regularly replicate designs worldwide. Distributing hardware via globally...reproduction of the structure in a similar way to the biological protocol, and assembly designs based around the...

...your application a whole week before the deadline. The day of submission, review your application with... their dream.  To help students focus on the application process, and simplify access to academic resources...Scrutinize every opportunity prior to starting an application to assess your eligibility, interest, and career... detailed. Beyond the resume, a cover letter communicates your interest in a specific program, and institution...professors, and mentors. Ask for specific guidance on grammatical errors, sentence structures, and overall format...February represent peak season for internship applications. Thus these noble individuals are inundated ... with a stellar letter of support.        5. Application submission  Set a calendar reminder for every...

...limited understanding of plant biology is a significant technical obstacle to plant synthetic biology. We ...biotechnologists face is public perception of genetic modification (GM) and synthetic biology. The mistrust and...emerging engineering technology. Monoculture and chemical spraying, mainstays of modern agriculture, are... by developing a plant biotechnology product specifically for consumer sales. Flowers that change color...a new conversation about GMOs.  Challenge #2: Technical obstacles to plant synthetic biology In addition...cannot build what we do not understand, and the complicated and unique metabolic processes of plants are ...creative problem solving like the Howard Hughes Medical Institute have begun to pivot towards plant biotechnology...

...mice at a time. Because mice are more genetically and physiologically similar to humans than many other model... of mice as model organisms is that they are genetically tractable. Mice can be easily manipulated with...also relatively straightforward to manipulate genetically. Scientists can make a new line of flies in only...reasons above but because they actually share many biological properties and processes with our own cells. ...important experimental model in the study of neurological diseases, such as Parkinson’s disease (PD). ... throughout their lifetime. Thus any genetic modification, such as expression of fluorescent protein or...embryonic development to adulthood. This ability to genetically manipulate and visualize cells and tissues allows...

...immunofluorescence? Generally speaking, IF is an application in which antibodies that have been joined to ...molecules, knowing that where you see fluorescence indicates your target’s location. Some other terms you might...ICC or a tissue in the case of IHC. IF is used specifically when we visualize the antibodies using fluorescence...bind to a single primary, resulting in signal amplification. Another reason is cost effectiveness. Generating...are not exclusive to IF, but they can have a significant impact on the outcome of your IF experiment. ...controls to show that your antibodies are binding specifically. For positive controls, aim to use samples that...antibodies you choose for your experiment will have a significant impact on the protocol you use and the ultimate...

... apply it in a clinical setting. CRISPR is often used for animal germline modification, to repair or add... important proof of concept for future preclinical and clinical work. Find Plasmids for Disease Research...exon skipping is a good option for DMD treatment. Clinical trials have used oligonucleotide exon skipping...estimated that exon skipping therapies would be applicable to 80% of DMD patients. Figure 1: Exon...that AAV-CRISPR does not cause obvious germline modification, an important finding given the controversy ...six months. Future directions and obstacles to clinical CRISPR editing Given the success of the CRISPR-mediated...researchers are very excited. This approach may also be applicable to a number of rare genetic diseases caused by...

... DNA – right? RNA performs almost all of its biological functions in our cells in the single strand form...generate dsRNA as part of their infection and/or replication process within the host. dsRNA viruses are inherently...generate dsRNA intermediates as part of the viral replication process. dsDNA viruses yield dsRNA through a ...machinery of the host. This pathway has been clinically exploited for antiviral purposes similar to that...sequence. Once a compatible sequence is found (typically RNA that the RNAi is designed against), RISC can...target mRNA. These RNA tools have been used in clinical therapy to target pathogenic proteins, as well...and insecticides, an advancement over previous chemical versions with toxicity to many organisms. ...

...Bárbara Pinho, a science communicator at the Portuguese science museum "Fábrica Centro Ciência Viva" in... these stromal cell modifications, stromal cells are also constantly communicating with a tumor, proving... is currently a science communicator at the Portuguese science museum "Fábrica Centro Ciência Viva" in...tumor recurrence and poor clinical outcome in human breast cancers." The American journal of pathology 174.6...reprograming influence tumors development? Being pathological active regions, microenvironments shape a tumor...Proficient in transportation, exosomes are tools of communication and provide malignancy to the microenvironment...interested in science journalism and writing about biomedical issues. Follow her on Twitter @BrbaraPinho6. ...

...detect mRNA from 10 bacteria of clinical significance and four clinically relevant host biomarkers. The ...microbiome with sequencing is vast, the ability to genetically alter microbiome members is limited and many ...at Columbia University developed a method for genetically altering gut microbiome members in their native...modular suite of plasmids (pGT) with various replicative origins that can be used for broad or narrow ... analyze gut microbiota and host biomarkers In clinical and diagnostic labs, deep sequencing and qPCR ...The lab validated their platform on clinical stool samples by comparison to qPCR and demonstrated that it...Central PMCID: PMC5963681.  Meyer, Richard. "Replication and conjugative mobilization of broad host-range...

...these types of bigger picture problems systematically and logically is as useful for finishing your thesis... but this skill is not just about reacting to technical issues. This skill also comes into play while ...defining problem solving skills as: One’s ability to logically think through a problem/task/situation, come up...Problem-solving overlaps heavily with other skills such as critical thinking, decision making, creativity, or data...might even be expected to write up a report to communicate your findings to your colleagues. So perhaps ... really are, while also practicing systematic, logical thinking.  If you are already a lab veteran at ... to troubleshoot; you probably did not just automatically come to a solution. Consider the thoughts that...

...through our lab using barcoded tubes and plates. The critical pieces of information for a given plasmid are ...plasmid sequence. Other information may also be critical to the maintenance and use of certain plasmids...access and use the materials they need, and avoid duplicating your effort. In addition, if you should choose...for the process. Always sequence verify Once the physical sample and its associated information are at Addgene...recover around 750 bp of clean sequence from a typical reaction, or about 250 amino acids worth, so make... for more tips on sequence analysis, plasmid verification, and using NCBI BLAST. Tips for high-throughput...comparable success rates and sequence quality, which is critical for the efficient handling of large numbers of...

...Red arrows indicate a conversion directly catalyzed by the base editor. Black arrows indicate natural cellular...editors (CBEs) and adenine base editors (ABEs) chemically modify target bases without breaking the DNA ...one-ring pyrimidine base. This is a far more complex chemical task, not a job for a single enzyme. Base ...editing works in general. Base editors use either catalytically dead Cas9 (dCas9) or a Cas9 nickase (nCas9) ... an engineered adenosine deaminase capable of chemically converting adenosine (A) to inosine (I, treated...converting the C base to uracil (U). This is the same chemical reaction performed by CBEs. However, most CBEs...improved editing in various cell types, although efficacy varied among target sequences. The researchers...

...plasmids available at Addgene. We will still periodically focus on specific CRISPR plasmid tools more ...editors Nme2Cas9 CRISPR interference in Candida albicans E. coli genome-wide inhibition library To expand...libraries also include 2,400 gRNAs without any significant hit across the E. coli genome as a negative control... control. Read the paper in Nature Communications Find the E. coli genome-wide CRISPR inhibition library...Homology-directed repair (HDR) can be used to make precise modification in genome editing. However, HDR efficiency is...efficiency up to 30-fold. Read the paper in Communications Biology Find the Cas9-PCV and PCV-Cas9 plasmids... fold and >3,800 fold in human cells with no significant reduction in editing efficiency. Read the paper...

...characters. A descriptive purpose focuses on the practical application of the plasmid and will help a scientist...species can be indicated in the “Species Other” column. Relevant Mutations Indicate the amino acid...usually very easy — our tech transfer team will communicate with your organization’s technology transfer ...name and match the name as it appears in its publication or as it is commonly known in your lab. Plasmid...reagent/antibody, unspecified, or other. If other, indicate type in “Vector Type Other” column. For example...or room temperature. Growth Strain Please indicate which growth strain Addgene should propagate your...For plasmids that contain the R6Kγ origin of replication, choose the Pir1 strain. If your plasmid cannot...

...genetic modifications into mouse embryos. But once you generate a growing colony of genetically engineered...epigenetic modifications. Special breeding cases Whether your mouse colony consists of genetically engineered...Cre-lox lines, and how to properly control for genetically engineered mice in your experiment. As you’ve...ve learned in Part 1, there are many types of genetically engineered mice: transgenic mice, knockin and...techniques are each useful for introducing one modification into the mouse genome, they are not commonly...parental mice In order to ensure that your genetic modification will be passed along to all of the offspring...getting a small piece of tissue from each mouse (typically via either a blood draw, tail clip, or toe clip...

... Molecular Biology (roughly a B average in the American system), in 2009 (at the beginning of the financial... this summer position as I felt I needed more practical lab experience to later gain a full time position...really enjoyed some aspects of the work, such as physically carrying out experiments like PCR and running...experience during research. Working abroad in rural Africa Once this position ended, I wanted to explore another...least 3 months experience working in a developing tropical country. This is why I set up a placement working...first aid. This really allowed me to see the practicalities of working in the field with the most basic...recession. As each advertised job received many applications, employers were requesting that only scientists...

...What’s behind these sights are an abundance of biological molecules like chromoproteins, fluorescent proteins...proteins and luminescence have been widely used in biological research for a long time, chromoproteins are ...just now being developed for a broad range of biological inquiries.  What is a chromoprotein and how ...photosynthetic activity. This still allows photosynthetically active wavelengths to reach zooxanthellae... sequencing are more definitive, but visual identification is a good place to start before taking on more...resources are limited or for bioart and teaching applications. Find a chromoprotein for your experiment! ...events: The plasmid carrying the desired genetic modification integrates into the genome (Figure 3). Here,...