...as mouse IgG2 or IgG4. This can be important for some applications. Assay use While secondary antibodies...much easier said than done! Each technique, and sometimes each individual assay, will need to be developed...attaching to a single primary antibody, it is sometimes assumed that the direct method is more quantitative...improve direct methods and avoid the indirect ones. Some Western blots, for example, use a primary antibody...

...panicked silence, I opened my mouth to panic-babble something about Western blots that made no sense, even in...options were to stop talking until I could present something coherent or to quickly string together all the...noteworthy and memorable and awful. So try to keep some perspective. Most people won't notice, or won't ... above to make a strategy that can help. Even something as simple as telling yourself, “I can stop and...

...needs to be empirically determined. In addition, some metal ion substrates, such as those containing Co2...to the metal ions once the protein is folded. Sometimes a tag placed on one end will be blocked during...not disrupted by denaturants (Hochuli, 1988). In some cases, proteins can be bound under denaturing conditions... Introduction to Antibodies.) Unfortunately, for some proteins there just aren’t any good antibodies available...

...are encoded by single stranded RNA (ssRNA), while some exist as dsRNA. Still others use dsDNA to store ...know the sequence of a virus, you could introduce some of its dsRNA intermediates into a healthy individual...Endogenous sources of duplexed RNA DNA:RNA hybrids Sometimes ssRNA just needs a buddy, and even DNA will suffice...documented in several autoimmune disorders, including some forms of lupus and rheumatoid arthritis. These ...

...so-called “small molecule degraders” recruit the proteasome to destroy targeted proteins. We previously developed...polyubiquitination of the target protein and proteasomal degradation. Remarkably, small molecule degraders...dTAG) system, to co-opt the endogenous ubiquitin proteasome machinery to degrade any protein. We are particularly...between the FKBP12F36V-fusion and the ubiquitin proteasome machinery through binding with cereblon, leading...

...itself: Ma et al. were interested in correcting an autosomal dominant mutation in MYBPC3 that leads to hypertrophic...term ‘editing.’ While repair using endogenous chromosomal DNA is appealing for heterozygous mutations, ...seems impossible with current tools, thus quieting some fears about the creation of “designer babies.” The... a ‘magic bullet’: its use will always come with some uncertainty. As biologists equipped to understand...

... structure? Helping to answer some of these questions, and posing some of their own are the following....   The news is not all bad, however - there are some signs that the tides may be turning. In 2013, the...- this time with the aim of finding solutions to some of the problems that were identified last year.  ...

...learn about the tools available for bacteria and some of the applications for which they’ve been used....Different Cas orthologues and subtypes function somewhat differently and can be more effective targeting...and CRISPRa using RNAP-Omega-dCas9 fusions. Here’s some others. SoxS based CRISPRa In the search for a CRISPR...Cytidine and adenine base editing in Streptomyces Some of the most industrially and clinically relevant...10.1016/j.cell.2013.02.022 Ronda C, Pedersen LE, Sommer MOA, Nielsen AT (2016) CRMAGE: CRISPR Optimized...

...resolution. Stories usually have a theme too, which is something not inherently captured by the STAR method. I ...interview. Each interview is an opportunity to learn something and then apply it to the next interview. Perhaps...underdressed, but read emails from HR carefully because sometimes companies have relaxed dress codes for interviews...questions after completing each section of your talk. Some video conferencing software allows participants ...

...Every few months we highlight some of the new plasmids, antibodies, and viral preps in the repository... it with an “assist peptide” that facilitates endosomal escape. Combining Cas9-PAGE with lentiviral sgRNA... achieved high editing frequency (nearly 100% in some cases) and minimal toxicity, including in clinically-relevant... interest, its dimeric nature can interfere with some applications. Several labs have been working to ...

... measuring locomotion. Mice may need to be given some time to adapt to a testing apparatus. General stress...adult animals, and not targeted to a specific area. Some other technical considerations for stereotaxic injections...via RNAi or gene editing/deletion using CRISPR. In some cases, a decrease of the expression efficiency can...depression. In upcoming blog posts, we’ll discuss some of these behavioral studies in detail and share ...

...genome. AAV instead relies mainly on episomal expression. Episomes are circular pieces of DNA that reside...expression limitations may be too constraining for some experiments, particularly in vivo ones. In those...& Kampmann, M. (2023). CRISPR screening by AAV episome-sequencing (CrAAVe-seq) is a highly scalable cell...

... training prior to conducting mouse experiments. Some of these techniques are what we call “beginner”,...injection into the sciatic nerve. Mice likely experience some muscle soreness after injection, like you might ...and restraint method, subcutaneous injections take some practice to get the coordination down. Subcutaneous...theoretically be performed anywhere on the body, somewhere large, flat, and easily accessible (like the back...

...Do you ever wonder about the origins of some of the common techniques or tools you use in the lab? Take...antibody developed almost 40 years ago? Well…kinda. In some cases, antibody providers are distributing monoclonal...tag have been generated in an attempt to overcome some of Myc1-9E10’s shortcomings. Wait, 9E10 isn’t the...sensitivity (Schüchner, et al. 2020). In their study, some of the newer Myc-tag antibodies appeared to be less...

...right Tn7 end sequences. Donor bacteria with a chromosomal copy of the RP4 transfer machinery mobilize these...and recipient bacteria that lack an integrated chromosomal copy of Mobile-CRISPRi. For firmicute gram positive...tri-parental mating. Donor bacteria contain a chromosomal copy of the RP4 transfer machinery, which mobilizes... 1 transfer in every ~100 to 1,000 colonies, but some bacteria, including E. faecalis and P. mirabilis...

...never worked with AAV, so naturally I had to package some virus for my first experiment. It was a bit intimidating...intimidating, but I had my lab’s protocols and some great co-workers to help me out. Even with these tools...definitely made my life easier!  In this post, I’ll share some AAV production, purification, and titration tips...to create the iodixanol gradient layers, and has some great pointers that while help you master iodixanol...

...that forms chromosomes. These long range DNA interactions are often detected with chromosome conformation...factors, can increase a gene’s transcription. Sometimes enhancers are located thousands of base pairs ...created for each genomic loci of interest. This is cumbersome and also makes it hard to compare results across...

...about? Flame logo: The flame that appears beside some of Addgene’s plasmids is an indication of popularity...section of our plasmid pages is relatively new, so some of our older plasmids may not have an entry in this...Ordering: Check the Price of course! The price of some items vary, for example, plasmid kits are avaliable...store plasmids that need extra stability, such as some of our viral vectors, in NEB Stable cells. Resource...

...how-to videos on YouTube long before he learned (somewhat successfully) how to text or use Facebook. However...However you choose to communicate your science, some people will love it and others will avoid it like ...proficiency.  What kind of content are you sharing?  Sometimes your subject matter will drive the kinds of communications...convey an entire story or context via still images. Some things cannot be photographed or need text to properly...

...and their associated ancestries. While there were some exciting relationships identified, 48% were found...differences in gene expression could explain why some ancestry groups were differentially dependent on...dependencies are undercounted by approximately 20%. In some cases, this may lead researchers to deprioritize...the bias  While their thorough analysis uncovered some concerning biases within widely used CRISPR screens...