...emission at 560nm. Shortly after the initial article describing the cloning of firefly luciferase was published...the plasmid, which is then introduced into the desired mammalian, plant or bacterial cells. Expression...stop codon, which is then transfected into the desired cells. A hybrid mRNA transcript containing luciferase...expression and more complex cell treatments can be designed to further explore the pathway. Which luciferase...

...but can be minimized with high powered research designs. Unanticipated discoveries Finally, there are good...rates, should be reported as a new discovery that deserves confirmation with new data and a new, preregistered...Guidelines; preregistration to increase clarity in study design and analysis; removing publication bias with Registered...Science, Dr. Mellor worked with citizen scientists to design and implement authentic scientific research and...

...trillion connections (synapses.)  In this post we’ll describe the AAV retrograde serotype (AAVrg) and how it... this can cause challenges for the experimental design as these vectors can exhibit spurious expression...efficiency can have a major impact on experimental design and interpretation of results. Take extra care ...References 1. Tervo, D. Gowanlock R., et al. "A designer AAV variant permits efficient retrograde access...

...Addgene provides the scientific community? Katz: The most important resource that Addgene provides is an...is continuing to look at providing other things besides plasmids to the scientific community through Addgene...exciting about the open science movement is that it provides interesting and unique challenges for intellectual...

...New work from Konermann et al. and Yan et al. describes new Cas13d enzymes that average only 2.8 kb in...Like other Cas13 enzymes, the Cas13d orthologs described in these papers can independently process their... high targeting efficiency, both of which are described below. Yan et al.: WYL-domain-containing accessory...produced very similar cleavage products even when guides were tiled across a target RNA, indicating that...

...Harvard University. In my previous two posts, I’ve described the fundamentals of how to work with Drosophila...Drosophila as an experimental model organism. I then described the Gal4/UAS system used by geneticists to study....e. there won’t be positioning effects). When designing experiments where you want to express multiple...wondering, “How do I know which of my flies have the desired integration?” Not to worry! In flies, phenotypic...

...purification is now a reality thanks to OptoNBs. Recently designed by the Toettcher Lab, OptoNBs are photoswitchable... put AsLOV2 because here it was less likely to destroy the nanobody’s function than if inserted into the...blue-light induced binding. The lab used this general design scheme to generate two more OptoNBs, one against...exciting because using light to control signaling provides greater spatiotemporal resolution than pharmacological...

...’s social media toolbox. Besides advice for using Twitter, this post provides suggestions for using LinkedIn...use Twitter as a scientist. While this post is designed with beginners in mind, there’s some great reminders...sheet from Science Media Savvy This tip sheet provides advice for tweeting and blogging as a scientist...

...target specific mRNA transcripts for degradation. Despite how easy these experiments are to physically set...to prevent the incorporation of non-specific nucleotides into the dsRNA leading to increased RNAi efficiency... Lab’s recent Nature Biotechnology paper, they describe TurboID and miniTurbo, which utilize non-toxic...switch to be expressed. In this case, the gene encodes the LacZ𝛼 protein. Once expressed, LacZ𝛼 complexes...

... NEB's Resource describing Dam and Dcm Methylases Discovery of DpnI Review describing dam-/dcm- E. coli...mutagenesis. During this process, incorporation of a desired mutation into your plasmid of interest by PCR generates...only the non-mutated and methylated template is destroyed leaving behind a pool of mutated plasmids which...

...cleaved precisely as predicted with no addition of nucleotides to the 5’ spacers. The 3’ ends were modified ...advantage over other methods that favor certain nucleotides at the 5’ end. PTG construct schematics. Scissors...the plasmids made by Xie et. al are specifically designed for plant expression, PTGs will likely be adaptable...worries, I've summarized the three strategies described in this post in the graphic below - including ...

... for over 10 years. Many successful scientists describe having a “posse” of mentors as one key to their...Ellen Daniell. In “Every Other Thursday,” Daniell describes a group of 7 women scientists who met every other...increases the diversity of input and perspectives Provides a good mechanism for accountability that makes...leadership and communication skills. The leader also provides materials such as pre-reads, videos or exercises...

...shown in the figure below. Their most successful design utilized Magnet photoswitchable proteins derived...architecture. Yu et al., 2020 FAST BphS from R. sphaeroides Split Cas9 Genome editing by NHEJ and HDR. Can...Cas9 Richter et al., 2016 tsRC9 RsLOV from R. sphaeroides Cas9, dCas9 Gene editing Zhou et al., 2018 ...al., 2016 CRISPR-plus Photocleavable ssDNA oligonucleotides Cas9, photocaged-gRNA Gene editing Manna...NPPOC-caged trimethoprim (TMP) Cas9 fused to destabilizing domains (Cas9-DHFR) Gene editing, transcription...

... as the OG (as the kids say) affinity reagents deserved to be celebrated, there are many other amazing...its thorns and every affinity reagent has its downsides. These reduced-in-size options often have reduced...°C and can refold correctly after denaturing.  Designed ankyrin repeat proteins, or DARPins, are genetically...a wide variety of targets, including proteins, peptides, amino acids, drugs, metal ions, and cells. They...

...molecular biology experiments was in the job description, but this skill is not just about reacting to...puzzling through your broader research questions and designing future experiments. Being able to work through...research non-academic careers, I always felt that descriptions of transferable skills seemed fuzzy or ill-defined...most likely cause, you implement an experiment designed to fix the problem. Then you assess whether your...

...in science – the time spent by the scientists designing and performing the experiments. This cost varies...quickly with the postdoc who’s really good at primer design – but first, you have to know what resources exist...well) plate! How much do repositories help?   Designing precise calculations for any of the numbers in...engagement with and use of repositories like Addgene provides large-scale savings. How large? Using Doench’s...

...and SMASh degron tagging vector! Destabilizing domains Destabilizing domains (DD) are akin to degron ...   Fig. 2 Tetracycline on/off promoters and modes of action Check out Addgene's Lentiviral Tet-on ...post-translational level, unlike most of the other mechanisms described. Fig. 3 dTag mechanism of protein degradation...

... plasmid maintenance during cell division. ccdB codes for the toxic protein (CcdB) that acts as a DNA ...death. ccdA, another gene found in the ccd operon, codes for the antitoxin protein (CcdA) that protects the...of ccdB, leading to positive identification of desired clones.   ...Becomes a powerful cloning tool... efficiently, as those that do not contain the desired insert should not grow.   Figure 2: Image...

...we would be remiss not to at least mention. Oligonucleotides allow for sensitive detection of low abundance...strong protein-protein interactions with other peptides, like an Fc-binding domain (Figure 1).  ...Strong protein-protein interactions with other peptides. Created with BioRender.com.     Many biologists... conjugation. But hopefully, this introduction provides a good base from which to dive deeper!References...

...another. If you are using barcodes (and you should be!) bear in mind that your barcodes are being applied to...containers, not to the contents. Tightly coupling barcodes and containers might not be such a bad idea, tightly...Resources on the Addgene Blog Learn about How We Use Barcodes in the Lab Get Tips on How to Keep a Lab Notebook...