...others.  Meditate. Meditating during the day is a great way to take a mental break, while meditating right...Organize your data and prepare publications or your dissertation Organize data that you already collected into...into figures for papers or presentations. Learn a new skill for analyzing and visualizing your data, such...needing to have collected data. Work on your dissertation, even if your defense isn’t for a while. Getting...Data Hub to help the research community. Set expectations for your productivity Set a routine and stick...

...  An MTA is simply a contract or agreement that states the terms of use for a tangible research material...MTAs.  Addgene does a lot to ensure that we can facilitate a fast and agile MTA process. This is important...deposits in our repository. From 2004 to 2023, we facilitated 784,795 MTAs with 8,798 organizations in 111 ...111 countries. From 2019 to 2023, we averaged facilitating 89,477 MTAs per year.  The MTA Process at Addgene...MTAs for plasmids in our repository. Instead, we facilitate the signing of MTAs by the Provider and Recipient...signature on any subsequent MTA for that plasmid facilitated by Addgene — basically, instead of having to ...shared reagents should be used. Setting clear expectations from the beginning makes the sharing process...

...track of computation work comes with new sets of challenges. One challenge with computational analyses... of biologists are finding themselves using computational approaches to analyze large data sets (myself...what works, and what does not work. Careful documentation will keep you on task and will prevent you from...especially since you’ll want to dig in to the computation, but they will serve you well as you perform ... the analyses: Goal: Before you begin, briefly state the goal of the analysis, including background on... you to perform this analysis. Having a clearly stated, specific goal for each analysis will help you ...

... cutting edge genomic technologies to map the mutations that are present, and use prior knowledge (data...One central bottleneck is that most recurrently mutated cancer genes are rare and most of the individual...characterized? Some have developed exciting computational algorithms, such as SIFT or PolyPhen to approach...well as reference cDNA clones mapping to well-annotated cellular pathways). It wasn’t clear whether gene-agnostic...High-throughput Phenotyping of Lung Cancer Somatic Mutations. Berger AH, Brooks AN, Wu X, et al. Cancer Cell...perform additional functional assays for these mutations and map other cellular functions and phenotypes...to build saturating libraries of all possible mutations in many major cancer genes. These should be used...

... homologous recombination to facilitate simple gene replacement/mutation. Yeast expression plasmids used...microorganisms, yeast have both a stable haploid and diploid state which is useful for genetic analysis, as well as...apparently independent of a known specific sequence) dictate the replication of these vectors. S. pombe plasmids...missing nutrient. Scientists can exploit these host mutations by including a copy of a functional gene which...This can complement ura4- S. pombe, but the complementation is weak. yes   LYS2 L-lysine yes - Grow on... S. pombe  no no *FOA in fission yeast induces mutation in the ura5+ gene in addition to ura4+. leu1...

... DIO (Double-floxed Inverse Orientation) or DO (Double-floxed Orientation)) is a powerful tool that provides...channelrhodopsins that, in turn, trigger neural excitation, whereas yellow light activates halorhodopsins...resulting fusion gene is inserted in the antisense orientation with respect to the promoter to prevent its expression...recognition sites (e.g., loxP and lox511) in opposite orientations. Since Cre does not cause recombination between...therefore locking the opsin into the correct orientation to be transcribed (Figure 2) (Sharma and Zhu,... arousal in mice previously subjected to steady-state general anesthesia (Taylor et al., 2016). These ...strains of Halobacterium halobium differing in pigmentation." Biochemical and biophysical research communications...

...: Summary of Rosella’s excitation and emission spectra. Component Excitation Emission pH Range DsRed.T3... inhibitor. While this method does provide a quantitative measure of autophagy, it only measures bulk ...final read out of this assay. This is also a quantitative measure of autophagy, but, like the protein ...DsRed.T3. See Table 1 for a summary of Rosella’s excitation and emission spectra. DsRed is the permanent ... 508 nm ~6.5 - 9 *Optimal sequential excitations wavelengths as determined by Rosado et al: see... T., & Klionsky, D. J. (2008). Chapter 3 The Quantitative Pho8Δ60 Assay of Nonspecific Autophagy. Methods...

...and methylated states. Thus, this system is a reliable tool in reporting methylation status of these sites...the headlines, with sotires like the potential devastation of the palm oil industry through epigenetic effects...grandchildren; in this study,epigenetic changes were statistically significantly associated with reduced life-span...syndrome is characterized by low muscle tone, short stature, cognitive disabilities, problem behaviors, and...epigenetics; techniques include chromatin immunoprecipitation, fluorescent in-situ hybridization, methylation-sensitive...methyllysine binding domain causes a structural change (facilitated by the flexible linker) that brings the fluorescent...

...and immunology. They can be used for qualitative and quantitative measurement of protein expression in...protein via fluorescence or chemiluminescence, precipitate the protein out of solution, or isolate cells...epitope of the target protein. However, small mutations in the DNA of the hybridoma can occur, known as...types or treatment conditions. They can be used quantitatively if you also load the gel with appropriate controls...media from cultured cells. ELISA can be used to quantitatively measure the amount of protein present. The ...ELISA (Enzyme-linked Immunosorbent Assay). Immunoprecipitation Sometimes, instead of just detecting the ...immunofluorescence), or by a colorimetric reaction facilitated by an enzymatic reaction. Figure 5: Immunofluorescence...

...assays include immunoprecipitation mass spectrometry (IP/MS) and chromatin immunoprecipitation (ChIP). In ...In IP/MS, a target protein is first immunoprecipitated - along with its associated protein complexes - ...tag” such that these tagged proteins can be immunoprecipitated using a standard tag-specific antibody (Jarvik...double-strand breaks (DSBs). DSB repair by HDR facilitates in-frame insertion of tag-encoding sequences ...efficiency across different stem cell types. This has facilitated easy generation of clonal knock-in lines that... design, remember to incorporate PAM-blocking mutations (e.g. convert the NGG PAM to NGC in your repair...2018), however, if you have any questions, do not hesitate to contact me at [email protected]. Keep ...

...2xNLS) and c-terminal (Cas9(C)-2xNLS) fragments facilitates target DNA cleavage upon co-expression and spontaneous...has been cumbersome. To overcome many of the limitations intrinsic to DNA-guided systems such as TAL effectors...or invert a specific chunk of DNA. However, a limitation to this strategy is that each "bit" of information...This leads to 2048 (211) possible combinations of states. Each of the integrases is supplied on its own ...more powerful LID, the authors recently used computational protein design, phage display and high throughput...system. How does it work? RUSH is an ingenious two-state assay based on the reversible interaction between...synthase pair is encoded on a single plasmid and facilitates the incorporation of a variety of bio-orthogonal...

...but unsure how best to prepare. Maybe you’re a presentation veteran, but have difficulty answering questions...what you would like to say to help build your presentation. For example, when thinking about discussing... Practice, practice, practice: Rehearse your presentation with people inside and outside your field. Your...and PI can provide feedback on your technical presentation, but those outside of your field can point out...you a chance to determine how to tailor your presentation so that it can be understood by your audience...It’s likely you won’t know every answer – don’t hesitate to be honest and tell your audience just that....attendees can get sucked in to thirty-minute-long presentations, which constrains the number of posters you ...

...the late 1980s, scientists even created a codon adaptation index (CAI) that is based on the codon usage ...codon bias. The idea is that scientists can make mutations throughout a gene of interest based on an organisms...) to name a few. This could affect synonymous mutations which have even been linked to some diseases. ...human genes contain a region where a synonymous mutation can be harmful (Sauna and Kimchi-Sarfaty, 2011...tabulated from international DNA sequence databases: status for the year 2000. Nucleic Acids Research 28:292... Understanding the contribution of synonymous mutations to human disease. Nat Rev Genet 12:683–691. https...10.1038/nrg3051 Sharp PM, Li W-H (1987) The codon adaptation index-a measure of directional synonymous codon...

...single protein, computationally described as a fusion of four Cas7 proteins and a putative Cas11-like protein...patient's specific mutations. With a drag and drop approach, however, the specific mutations wouldn’t matter...evolution, were also working on the project. Using computational observations, Koonin and Makarova found that...which can be caused by thousands of different mutations, all found in the same gene. If we only have small...matter – the mutated copy could be replaced by simply dragging and dropping in a functional copy of the entire...

...fluorescence that is generated allows for qualitative and quantitative measurement of processes including RNA...photobleaches, DFHBI can only be used for qualitative, but not quantitative, studies. Additionally, Spinach, the... In the search for something more stable for quantitative imaging, Ming Hammond’s group created Spinach2...photostable than Spinach and Broccoli, and useful for quantitative studies. Specifically, Corn was developed to... and both TO1-biotin and TO3-biotin contain an acetate modification that reduces nonspecific binding with...

...biotin ligase, BirA*, with a site mutation (R11G). This site mutation destabilizes the ligase, allowing...the E. coli version. It also contains a single mutation (R40G) in a conserved residue in the biotin catalytic...ascorbate peroxidase for proximity labeling A major limitation of APEX was its low sensitivity. Sensitivity ... two mutants, the 35 kD TurboID, which has 15 mutations compared to BirA and the 28kD miniTurbo, which...which has the N-terminus domain deleted and 13 mutations compared to wild type BirA. TurboID and miniTurbo...interactions Split-BioID combines a protein fragment complementation (PCA) technique with proximity labelling to...

...template to create mutations HDR donors can contain insertable sequences or specific mutations, along with homologous...sequence at the DSB, or it can introduce specific mutations into the damaged DNA if these modifications exist...HDR efficiency for more details on how "Cut-to-Mutation distance" affects editing efficiency. One important... cutting and repairing your target DNA until a mutation is introduced disrupting one or both of these ...sequences. To prevent re-cutting and the resulting mutations after an HDR event, ensure the gRNA or PAM site...off-target effects. A technique that pushes the size limitations of ssODNs is Easi-CRISPR (Quadros et al., 2017...

... introduce naturally occurring cancer-causing mutations into the endogenous allele of the STAG2 tumor ... introducing non-sequence specific frameshift mutations through aberrant double strand break repair, it...knockins, it is not always ideal. Because of these limitations, the AAV approach is best suited for tissue culture...epitope tags, or the introduction of dominant mutations of a gene into the endogenous allele in cultured...template to mouse zygotes and thereby correct a mutation that causes Hemophilia B. Unfortunately, work ...applications that require large gene fusions. The current state-of-the-art in combining the AAV and CRISPR-based...segregation in human cells harboring tumor-derived mutations in STAG2." PLoS genetics 12.2 (2016): e1005865...

...within this kit contain a variety of promoters to facilitate gene expression in bacteria, yeast, mammalian...His, Avi, and GFP) as well as protease sites to facilitate protein purification. We hope this kit will allow...important microalgae: intrinsic suicide switch implementation in cyanobacterium Synechocystis sp. PCC 6803...Furthermore, some plasmids contain peptides to facilitate protein folding and/or protease sites to remove...The gene set was carefully chosen to perform statistically relevant benchmarking of cell-free expression...By counting DNA sequence reads, FREQ-Seq can quantitatively determine allele frequencies across timepoints...selective pressures: Miller, Corwin, et al. "Adaptation of Enterococcus faecalis to daptomycin reveals...

...purified GFP could be imaged in the green state after excitation at 488 nm, photoconverted with 405 nm light... light, and then captured in the red state after excitation at 561 nm. This imaging process worked well...S65T GFP, and no red color was observed after excitation at 488 nm, indicating that spontaneous conversion...al., Sci Rep. 2015 CC BY 4.0 Applications and limitations of GFP photoconversion One concern with photoconverting...