...and genome modifications to reporter vectors (e.g. pBV-Luc) and gene silencing (e.g. pLKO.1 - TRC cloning... (e.g. 6xHis containing pDEST-HisMBP). After purification the protein can be released from the resin and...types of yeast expression vectors. Consider your applications and experimental design. You can create a new...more traditional approach of an autonomously replicating vector that carries either a wild-type version...a variety of other types of vectors for most applications used to study gene function. From making transgenic...

...could be kept frozen after collection. Figure 1: Paul included a voltmeter on top of the battery box...under harsh conditions. One example application from the publication is the removal of unwanted cDNA derived...the van der Oost lab. Phylogenetic comparison indicates that like GeoCas9 and ThermoCas9, IgnaviCas9 is...a thermostable Cas9 be useful? It can enable applications where reagents must be stored at elevated temperatures...

...year and happy reading! Top 10 blog posts of 2015 1. CRISPR 101: Homology Directed Repair Addgenie Chari...overjoyed to provide you with these fantastic educational resources and will be sure to keep our eyes open...used to generate inversions, deletions, and translocations. It also provides researchers with many different...repository use Cas9 to cut DNA, Addgene Science Communications Intern Mary Gearing explains how Cas9 homolog...make it easier to make gene edits at new genomic locations. 6. CRISPR 101: Non-homologous End Joining So ...

...help you decide if a master’s might be for you. 1. Are you interested in master’s-level jobs? Ultimately...technical know-how, and helped develop my science communication skills. Ultimately, my degree prepared me to...technical or product support specialist,” “field application scientist,” or “scientific or technical writer...for PhD applicants since many will also accept applications from master’s candidates with experience.  Use...

...specific DNA sequence in only 1 min. Using these lentiviral plasmids, catalytically inactive Cas9 (dSpyCas9)...makes them more adaptable to a broad range of applications. Recently, the Spiess lab developed a set of... with specific genomic loci. He made use of catalytically inactive dCas9 by fusing it to an engineered...

... to just that particular assay.      Fig. 1: A native protein (left) is denatured, allowing the...Conformation changes usually fall into one of two categories: ones induced by the researcher and ones that...naturally, by binding to other proteins, changing locations, or even just as part of performing a specific...it a transmembrane protein, or is it know to translocate or fold in certain conditions? It can help to...

...evidence through education.  My only criticisms of what I saw are that we, as a society, need to: 1) use more...impartial. The best science happens when scientists have 1) the courage to make mistakes and 2) the integrity...also a science communicator, UBC Public Scholar, and a member of the Vancouver SciCATs. Follow her on ...researchers and supporters of science marched to advocate for scientist participation in government, evidence...promote help motivate you to spend a little time advocating for science and getting yourself out there for...April because devaluing science has alarming implications for our public policies—many of which impact...scientist. I’ve worked tirelessly to get the publications, grant funding, and teaching experience that...

...targeting MNase to antibody bound proteins (Figure 1). Importantly, since Protein A has weaker affinity...interactions for subsequent sheering of DNA by sonification. Though it helps capture weaker or more transient...change offers a significant improvement and simplification of the original technique, specifically in ...

...updated version of BRET called BRET2. Figure 1: TRUPATH measures heterotrimeric G protein dissociation...Kadowaki S, Haga T, Takaesu H, Mitaku S (2002) Identification of G protein-coupled receptor genes from the...biosensors Resources on Addgene.org Browse the catalog of fluorescent protein plasmids Find biosensors...

...can also rank the plasmids by abundance, scaled to 1, to create a “cumulative wealth distribution” curve...would be a good time to talk about the amplification and verification processes we use to ensure high-quality...your own experiments. Library amplification 101 Library amplification seems straightforward: E. coli ...transformed with the library DNA, they grow and replicate, and then their DNA is harvested, much like one...library has its own depositor transformation and amplification protocol, but the overall workflow is similar...ll need to remember when undertaking library amplification and sequencing. Maintain representation of the...electrocompetent Stbl4 cells when performing amplifications, and we use electroporation to ensure a good...

...net negative charge with 4 negatively charged and 1 positively charged amino acids. Myc tags are frequently...(stay tuned for our affinity purification tag blog!).   Applications of epitope tags   The...success rates across all applications. You can find FLAG antibodies on Addgene’s catalog including a mouse monoclonal...protein purification is not ideal with Myc, as low pH conditions are required during the purification, which...experienced. Finding an antibody that works for your application, specifically detects your protein, is species...and have been widely used across species and applications. In this blog, we will review the most common...that are bound by antibodies for the purpose of purification or detection of the tagged protein. These peptides...

...the brain where DA quantitation is desired (Figure 1). The probe perfuses liquid, which causes the passive...at a certain frequency, and this frequency can indicate behavioral state, for example, if someone is awake...

...into three main effector functions.  Figure 1: Fc effector functions in an immune response to a ...determination, don’t generally affect antibody applications like ELISAs or Western blots, they are key in...

...motion videos is very time consuming; producing a 1-2 minute video takes an entire week of our free time...Research 5 Reasons to Use Reddit for Science Communication 10 Ways to Share Your Science Resources on ...

... Table 1: New ready-to-use viral vectors in the Addgene repository  Name Serotype Category Depositor...

...materials to the research community. References 1. Coroadinha, A. S., et al. "Effect of osmotic pressure...AAV yields. Given the wide range of downstream applications scientists rely on our AAV preparations for,...

...enables the high-resolution tracking of more than 1 million cancer cells under drug treatment.” Stegmeier... interference. In the first two months since publication, the lab got over 100 requests for the plasmids...

...References 1. Huang, Ally, et al. "BioBits™ Explorer: A modular synthetic biology education kit." Science...plasmids and their potential educational uses. We believe that biology education is something that should ... expensive tiny amounts of reagents are, how complicated (and again, expensive) the different machines...desire to create a set of low-cost, easy-to-use educational biology activities – that any school could afford...proteins without the need to handle a living, replicating organism. Furthermore, cell free systems can ...in adapting your research ideas into hands-on educational activities using the BioBits platform, please...BioBits™ Bright: A fluorescent synthetic biology education kit." Science Advances 4.8 (2018): eaat5107. PubMed...

...difficult to modify in the past.   Figure 1: Identification and enrichment use of XMAS-TREE. a) Cells ...  After identification of ABE activity, XMAS-TREE can also be used to purify these modifications and work...ABE-targeted cells By Ellie Adams Single base modifications through ABE or CBE's have the potential ability...point mutations. They allow single-nucleotide modifications to edit DNA without requiring a double-stranded...mCherry and GFP expression to check for ABE’s. b) Quantification of base-editing efficiency using XMAS-TREE-...methods at various genomic loci in four different categories.     Brookhouser, N., et al. BMC Biol 18...MYC-dependent cancer cell lines. It can be applied for identification of essential MYC binding sites in a wide range...

...vesicles. Okada CY, Rechsteiner M. Cell. 1982 May;29(1):33-41. PubMed. Protein transduction: unrestricted...protein-only transduction method and its potential applications to CRISPR/Cas9 genome editing.   Protein-only...the HIV protein Tat. Although CPP fusions can translocate a cell membrane, the presence of the CPP may ...that components in the buffer used for protein purification must be helping the untagged protein enter cells...Diego D'Astolfo. CRISPR/Cas9: a potential application of iTOP Having tested iTOP using Cre-lox recombination...showed up to 70% resistance to diphtheria toxin, indicating a 70% frequency of biallelic mutations. Importantly...Importantly, hESCs retained their stemness, indicating that iTOP may be valuable for CRISPR/Cas9 stem cell...