...study its function is one of the most popular applications of fluorescent proteins. These fusion proteins... from Aequorea victoria and its variants are genetically encoded fluorescent probes. One of the limitations...motifs that are necessary for post-translational modifications. Despite these potential issues, GFP has been...ultracentrifugation or gel filtration is the most common analytical method. However, these may not reflect the situation...situation in cells. Therefore, the localization of critical fusions (e.g. tubulin or connexin) has been used...linker sequence short In the early days of GFP applications, many were concerned with steric hindrance by...fluorescent protein and protein of interest (POI). To indicate the sequence, people often use terminology like...

...components of the cytoskeleton. They provide cells with physical structure, separate chromosomes during mitosis...filopodia formation, cell contraction, or other morphological changes, depending on the specific Rho GTPase...RhoGEF is a great way to stimulate and study these physical processes. Joachim Goedhart and colleagues in ... binding of the plasma membrane. In previous publications, they described tools to control formation of...the effect to a specific time or location can be critical when, say, the gene is essential during development...isn’t powerful enough, what about altering the chemical composition of the membrane itself? The Baskin...Baskin Lab previously published OptoPLD, an optogenetically controlled phospholipase D (PLD). Now they present...

...cryogenic electron microscopy (cryo-EM) protocols typically require high concentration and purity of the target... (Figure 1). This enrichment streamlines the purification and minimizes sample loss while also supporting...chaperone NPM2, but the technique should be widely applicable to other targets. The modular system of linkers...scFvs for other targets or could be used for applications beyond cryo-EM.   Find plasmids for MagIC-Cryo-EM...interact with the air-water interface (AWI) before vitrification, they can be completely denaturated (left) or...right). B) Protein samples protected from AWI by helical LEAs. C-E) cryo-EM single-particle analysis of ...orientations, this could be overcome with methods such as chemical crosslinking or tilted-stage imaging. A truncated...

...people who are data oriented, scientifically minded, and able to communicate complex information - all traits... incredibly important to the stability of the biological research enterprise going forward, in this post...the FOR website, the FOR Boston 2015 organizers indicate that the main question they wanted to focus on...Publishing Panel brought in editors from a variety of publications as well as young PIs and all gave advice on ...blogging, write reviews, or write for a university publication, improving your general writing skills will help...academic publishing goals and make you a better communicator overall. Don’t Write in a Vacuum Your research...cognizant of this during my PhD. Submitting a publication doesn’t have to be like throwing it into a black...

...non-viral methods including both chemical and physical methods in the lab. Chemicals like calcium phosphate and...Scientists can also use electroporation, the application of an electrical charge to cells, to increase cell membrane...take up DNA, they can be made competent through chemical manipulation in the lab. This is commonly done... properties. The phagemids are packaged into replication-incompetent phage particles with assistance from... found on an episome, a piece of DNA that can replicate on its own or be integrated within a bacterial...easily cross the negatively charged cell membrane. Physical methods such as electroporation or microinjection...cells. Electroporation on the other hand uses electrical pulses to create transient pores in the cell ...

...potential applications. Challenges of existing mutagenesis methods Several continuous diversification methods... challenging for users to achieve targeted diversification of all nucleotides within a defined region ...Chang Lab have created methods that introduce diversification by targeting a site of interest with dCas9 ...level of variation that can be introduced.   Diversification is limited due to the integration of oligonucleotide...increasing the standard mutation rate seen during DNA replication by 120-fold over background. Due to its modular...only ~2-fold over nCas9 levels. Two sets of modifications were also made to PolI3M to increase the mutation...more than one mutation near the target site. Applications of EvolvR The Dueber Lab used EvolvR to discover...

... given prokaryote typically has genes encoding one or a few restriction modification systems containing...methylation sites are not specifically associated with any restriction modification systems, their sequences... one half of naturally occurring restriction modification systems that prokaryotes use to protect themselves... phage for instance). Aside from restriction modification systems, DNA methylation also plays an integral...integral role in regulating genome replication, repairing mismatched basepairs or small indels that occur...methylases) are independent from the restriction modification systems, yet can still affect whether certain... These specialized E. coli strains have been specifically engineered to be Dam and Dcm methylase-deficient...

...tools from across the biological spectrum to learn more about the brain and neurological disease. I was surprised...and, like many biological fields the number potential of experiments enabled by physical, software, and...the things that popped out to me as a novice and indicate why other neuroscience newbies might want to get...brain functions and vice versa. Even within the biological sciences, there’s plenty of interaction between... microbial DNA they all seem to require some mathematical know-how for analysis. If you’re looking to ...presentation area, I honed in on projects using biological tools. In addition to the researcher using sequencing...but are afraid you don’t have the right kind of biological expertise to get into the field, there’s likely...

...of DNA that can potentially complicate results of a downstream application such as quantitative PCR to...sulfate on the activity of ribonuclease A. Biochimica et Biophysica Acta (BBA) - Enzymology 341:484–488. https...easier. Recently, we published a blog on DNA purification without a kit that outlined several reasons ...take care whenever doing any type of DNA or RNA purification to avoid contamination, take extra care when..., that isn’t the case for RNA extraction and purification. There is one simple method that works, and ... within cells or tissues- they don’t actually chemically stabilize the RNA molecules (Allewell and Sarma... MacDonald RJ, Rutter WJ (1979) Isolation of biologically active ribonucleic acid from sources enriched...

...suggesting that QKI is a host protein critical to Zika replication. Human genetic disease. RaPID can also...methods Not in a native context. While native purification preserves RNA structure and naturally occurring...present in the cell, it can also introduce non-physiological RNA-protein interactions due to sample processing...t be used to study endogenous RNAs at their physiological expression levels; however, knock-in of BoxB...used the Contamination Repository for Affinity Purification (CRAPome), to identify highly biotinylated background...research groups that performed BirA*-based affinity purification mass spec experiments. These background proteins...subtracted from Ramanathan et al.’s RaPID datasets. Applications of RaPID RaPID sounds like a nifty technique...

...regulation thats as sophisticated as a natural network. We could build a material as sophisticated as wood. To...to be able to design new materials and chemicals as sophisticated as those that living cells and animals...Technology has been “working on new experimental and theoretical methods to push the scale of genetic engineering...for major advances in agriculture, materials, chemicals, and medicine. As they’ve gone along, Voigt’s ... make a very large number of regulators. That basically means larger genetic circuits that could conceivably... perspective and you can layer them for more complicated functions. Those things are easy with CRISPR.... carbons and oxygens. Nothing too much more sophisticated. There are also proteins, like antibodies. But...

...responsibilities? Is the work (topic and/or technical aspects) applicable to industry if this path interests you...honestly evaluate your own style of work and communication before making this important decision. I’ll ...Prepare by going sleuthing on the lab website, publication records, LinkedIn, ResearchGate or Twitter. Be...the appropriate first authorship position on a publication of her project and made another lab member first...alumni, don’t ever disregard warning signs of unethical behavior, harassment or bullying. Even with the... including preventing access to opportunities, physical or social isolation, withholding necessary information...connections such as serving on Boards? Does the publication record show evidence of industry collaboration...

...primate. Likewise, in vitro efficacy doesn’t always translate to in vivo efficacy. You’ll therefore need to...Using AAV vectors in complex biological systems can be tricky at best, and downright infuriating at worst...site, or when using chemogenetic tools, where modification of cellular activity outside of your experiment... required is inversely related to transduction efficacy: the higher the efficiency, the lower the titer.... A. (2023). Various AAV Serotypes and Their Applications in Gene Therapy: An Overview. Cells, 12(5), ...Approaches and Further Perspectives of AAV Tropism Modification. Pharmaceutics, 13(5), 750. https://doi.org/...

...additional origins of replication compared to monomers, they are also able to replicate at a higher frequency...expected fragments for that plasmid. This phenomenon indicates the presence of a multimer.   Fig. 1:...the right, with the entire monomeric sequence duplicated in tandem, resulting in a plasmid that is twice...that affect the frequency of multimerization. Theoretically, as long as all the plasmid features remain ...other methods below should help in multimer identification.   Fig. 2: Gel image of Plasmid 61564...within a plasmid and, therefore, allow for the identification of plasmid multimers. Capillary Gel Electrophoresis...multimerization. One study by Bacolla et al. (2011) indicated that strain HB101 produced the highest yield of...

...— prime editing typically needs to be optimized for each application, and the efficacy varies widely —... the indicated tool. ~ reflects an innovation that may optionally be combined with the indicated tool....also more precise. Base editors, for example, typically edit all the target bases within the base editing... target nucleotides are positioned within the canonical base editing window, base editing has higher efficiency...products to be indels while the efficiency of HDR is typically less than 10%. In contrast, the original paper...editing variants Four years after their initial publication, the Liu lab returned to the reverse transcriptase...C-terminal end of PEmax to produce PE7. PE7 significantly improves editing efficiencies compared to PEmax...

...classrooms, the laboratory experience is focused on classical embryology techniques such as basic observation... in situ hybridization experiments. A recent publication [1] from the Sánchez lab describes the plasmids... manipulation. The technique involves using synthetically made RNA (riboprobes) with modified nucleotides... added to the tissue. The last step involves a chemical reaction to stain the cells that contain this ...easily generate off of a circular plasmid via PCR. Typically, to make these constructs, developmental biologists...the deposit) are described in the Sánchez lab publication [1] so that the students performing these experiments...target mRNA and silences its expression. More specifically, during the process of RNAi, an enzyme dubbed...

...human subjects or clinical trials Acknowledgement in recipient’s future publications relating to the ...results, modifications, and inventions created by the Recipient Right to file patent applications on modifications... about intellectual property rights, pending publications, and commercial use often come up. An MTA can...pride in facilitating the exchange of not just the physical samples, but also the legal documentation that... Addgene has adopted the terms of the Uniform Biological Material Transfer Agreement (UBMTA) for its material...have opted into our eMTA system can even sign electronically or choose to auto-approve MTAs. Whatever your...

...spatial and hierarchical organization of chromatin interaction networks  A second application for CAPTURE...created a dCas9-based CAPTURE (CRISPR Affinity Purification in situ of Regulatory Elements) method. The ...at once (Liu et al., 2020). CRISPR Affinity Purification in situ of Regulatory Elements: The original...but close together in terms of where they are physically located. DNA is then sheared into pieces small... original CAPTURE and CAPTURE 2.0 methods.   Applications of CAPTURE 2.0 CAPTURE 2.0’s increased rate ...identified with the multiplexed CAPTURE 2.0 largely replicated interactions previously identified with the original...CAPTURE 2.0 is the construction of spatial and hierarchical chromatin interaction networks. Think of these...

...setting—especially if job applications require teaching experience or high numbers of publications, which may take...their funding. Communication: In both industry and academia, you will have to communicate your science ...can be long—5 years or more, especially in the biological sciences—whereas industry postdocs have a set...course, you might have less time to explore scientifically since you can’t afford to spend too much time...Unless your academic lab is closely involved with clinical trials or has ties to industry, the drug discovery...people involved is staggering. The process is complicated. If you’re interested in the drug discovery process... proprietary compounds or information. Where publication is expected, the process is similar to academia...

... as a helicase. Following recruitment by Cascade, Cas3 nicks the non-target strand. The helicase domain... Cas effectors to carry out genomic editing. Specifically, these types of CRISPR systems work using a ...recruit Cas3, which carries out the brunt of the mechanical work for editing. Class 1 CRISPR systems make...After these initial steps, Cas3 can perform its helicase and nuclease functions simultaneously, by continuously...debilitating disease! This is, of course, only hypothetical at this point. More time and energy must be ...Mashimo, T. (2022). Genome editing technology and applications with the type I CRISPR system. Gene and Genome...unidirectional genome editing in human cells. Nature Communications, 10(1). https://doi.org/10.1038/s41467-019-...