...protein is up or downregulated, examine a protein’s stability or post-translational modifications, or study ...A and Protein G, which bind to the fragment crystallizable region (Fc) of antibodies with a high affinity...conformation. Frequently, the antibodies used for a standard, denaturing Western blot will not be suitable ...interest, such as a sample transiently transfected or stably infected with the protein of interest and a sample..., Hjelm K, Sjöquist J (1972) Protein a from Staphylococcus aureus. Its isolation by affinity chromatography...

...these different transcriptional start sites in order to better understand isoform-specific gene regulation...each gene isoform has a distinct transcriptional start site. Because CRISPRa can be targeted anywhere in... Bdnf gene possesses 9 distinct transcriptional start sites, all of which yield the same BDNF protein....downstream of the TUJ1 coding sequence. ES cells that stably express the dCas9-Suntag activator were transduced...

...longer shelf life and the development of disease-resistant crops to increase agricultural yield (Wang et ... experiment. CRISPR components can be expressed stably or transiently depending on the delivery method...be delivered and expressed transiently using a standard detergent, Polyethylene Glycol (PEG), although...Y, Liu Y-G, Zhao K (2016) Enhanced Rice Blast Resistance by CRISPR/Cas9-Targeted Mutagenesis of the ERF...homoeoalleles in hexaploid bread wheat confers heritable resistance to powdery mildew. Nat Biotechnol 32:947–951 ...

...be used with Type IIS restriction enzymes and standardized primers to clone your gene of interest into ...clone your gene of interest. Translation of the standardized leader peptide prevents secondary structures...The gene set was carefully chosen to perform statistically relevant benchmarking of cell-free expression... daptomycin reveals an ordered progression to resistance." Antimicrobial agents and chemotherapy 57.11...effects during adaptive evolution of antibiotic resistance." Proceedings of the National Academy of Sciences...

...promoters. Others are only active under specific circumstances. In a previous post, we discussed inducible ...which can be switched from a default OFF to an ON state, and how you might use these in your research. Today...which can be switched from a default ON to an OFF state, as well as repressible binary systems. In recent...GAL4 gene downstream of a tissue- or developmental stage-specific promoter and design a UAS reporter construct...QUAS promoter is less leaky than UAS in the basal state, and co-expression of the inducer QF can increase...

...alleles in a variety of species, including the standard laboratory mouse. It’s also made targeted insertions...protein knock-in, suffer from low efficiency when a standard dsDNA repair template is used. The original design...Easi-CRISPR efficiency. Easi-CRISPR is compatible with standard microinjection and electroporation methods using...editing components to the germline. In contrast to standard microinjection, i-GONAD is performed directly ...Gurumurthy, Masato Ohtsuka, and Rolen M. Quadros for assistance with editing this blog post. References Yang,...

...videos for you to level up in the lab, and we’ve started collecting some of our favorite (moving) images... You’ll be able to use these GIFs on Twitter, Instagram, TikTok and more. Just type “addgene” into the...looking for that perfect GIF for your TikTok or Instagram story, we also have stickers to help express your...check out everything we’ve uploaded so far, and start sharing the science love today!        ...

... critical systems that cannot be removed or substantially reduced in an attempt to save energy. The transition...renewable electricity is certainly not the only sustainability effort we’ve implemented at Addgene. The kitchen...check out Leila Haery’s blog series, “Bringing Sustainable Practices to the Lab.”  Whether participating...blog Subscribe to the Addgene blog Read our sustainability in the lab blog posts Read another blog post...

...transfection in: 15 cm dishes OR CF5 stacks Select how many dishes or stacks you plan to transfect: 1 to ...following μg of each plasmid needed per dish or stack. Calculations are based on using 28 μg of total ...) per 15 cm dish or 2500 μg of plasmids per CF5 stack. Total ug of each plasmid needed for the whole experiment...

...million bacterial stabs take up? If 81 vials fit into one freezer box, one million stabs would take up over... community (check out the infographic below for stats on our plasmid deposits and requests). Over the ...needs of the scientific community. For example, we started a new service distributing ready-to-use viral preps...

...both brain function and dysfunction. First, let’s start with the basics: What is optogenetics? Optogenetics...Controlling neurons with light In 2005, a team from Stanford University, led by neurobiologist Karl Deisseroth... central nervous system due to their relatively stable extra-chromosomally long-term expression and low... arousal in mice previously subjected to steady-state general anesthesia (Taylor et al., 2016). These ...therefore, an ideal partner to optogenetics to understand the cellular and molecular mechanisms of the ...

...We often think of DNA as inert. It generally stays put, which makes it easy to locate in a genome. But... in bacteria, transposons can move antibiotic resistance genes between plasmids or from plasmids to the...piggyBac is biased to integrate at transcription start sites, CpG islands, and DNaseI hypersensitivity ...Tol2 also prefers to integrate at transcription start sites, CpG islands, and DNaseI hypersensitivity ... of RNA guided transposons in bacteria sets the stage for combining the super powers of CRISPR and transposons...

...education content. This work exemplifies our long-standing commitment to be responsive to the community’s...environment. In particular, we are working to understand what challenges remain in materials sharing, ...innovation around the world. As always, we seek to understand our role in opening up science to help create...therefore embarking on a strategic refresh to understand this new world and our place in it. This period...

... the bioluminescent properties observed in the crystal jellyfish Aequorea victoria (Figure 1) in molecular...the Pacific Ocean, off of the coast of Washington state. Using these samples, he was able to isolate two... time. After Chalfie’s work, scientists really started to take notice of GFP, and began to realize its...resulted in much of our current mechanistic understanding of GFP’s function. His lab described the protein...

...like I understand the science but my prior HR experience at Harvard Medical School means I understand the... like “How do you start to bring people back to on-site work?” and “How do you start to expand operations...

...addition to learning how to be a scientist. Why not stack the deck in your favor? Look for opportunities to... amazing students at Washington University who started the BALSA group, a nonprofit that provides short-term...stick to timelines Be a mentor in a formal program Start something (a journal, a science lunch club, a biotech...by Peggy Klaus Looking for a mentor or want to start a peer mentoring group? "What Makes a Good Mentor...

...other name would…probably still edit genomes. Understanding the different types of CRISPR systems can be...will help you gain a deeper appreciation and understanding of CRISPR, and perhaps provide insights and ...main aspects of each class and type are easy to understand. Classes are based on the main Cas effector; ...highly unlikely that you will have to know and understand all the minute differences between the CRISPR...Makarova, K. S., Wolf, Y. I., Alkhnbashi, O. S., Costa, F., Shah, S. A., Saunders, S. J., Barrangou, R....

...cellular milieu. It is, therefore, critical to understand how proteins interact with one another (or with...tagged proteins can be immunoprecipitated using a standard tag-specific antibody (Jarvik and Telmer 1998)...Figure 1). In the first step of optimization, we started off by using custom synthetic two-part cr/tracrRNAs...and then analyzed for tagging efficiency using standard immunocytochemistry (ICC) assays.  Figure 1: ...a crRNA cutting 8-15 bp upstream of your gene’s Start codon will be suitable). For N-terminus tagging,...

...meet everyone but this never happened. I actually started at a later date and worked entirely virtually. ... I was able to express my visions for what my Instagram posts would look like. While working remotely ...continuing Addgene’s Meet an Addgenie Series on Instagram. I interviewed fellow Addgenies about their role... Some of recent Meet an Addgenie posts on Instagram! From this project, I observed what it was ...

...requires a substantial amount of properly folded, relatively pure protein, but getting to this stage is often...Additional tags may be used to improve protein stability and solubility. Determining the best way to express...Ni-NTA resin. MBP improves protein solubility and stability. TRX and signal peptide tags help proteins with...