...plasmid such as pX330. This cloning strategy not only makes it easy to create a single gRNA-expressing plasmid...clone up to 7 gRNAs into one destination vector, making multiplexing easy.  Disadvantages of Golden Gate...TAL effector-based constructs for DNA targeting. Cermak T, Doyle EL, Christian M, Wang L, Zhang Y, Schmidt...

...Requiring an additional plasmid for transfection can make these optimization experiments a bit more challenging...experimental volumes accordingly. While you may need to make some final adjustments, this is generally a good...additional resources. References Kumar M, Keller B, Makalou N, Sutton RE. Systematic determination of the packaging...

...vehicles for genetic information. Therefore they make excellent tools for genetic engineering. There are...vectors when used in experiments. Retroviral vectors make up a large toolbox used by researchers for (among...shRNA or miRNA. The ever expanding CRISPR field makes wide use of retroviral delivery systems, especially...

...spatiotemporal control or an “on/off switch.” This makes them great tools for providing a global overview... of the bait protein will be biotinylated. This makes it’s impossible to distinguish proteins that directly... proteins. This extended incubation period could make it difficult to detect proteins with short half-...

...it also helps you to hone your writing skills and make a name for yourself. Pick a topic that is close ...’t you record a podcast or put some effort into making a video? Use your unique talent to communicate ...Since then, I have tried many things: photography, making videos, presenting in pubs and at science festivals...

...proteins had to be re-engineered for each gene, making wide-scale gene activation seem next to impossible...activation compared to other CRISPR activators, making it a popular method for gene activation experiments...design (SunTag). This streamlines its delivery, making it a common choice for CRISPR activation. Newer...

...crucial if we want to make sure that everyone can be informed and empowered to make their own decisions ...posts, lesson plans, or information briefs for policymakers. The rapidly changing landscape of genetics ...

...precipitates that permanently stain the membrane, making that section of the blot unusable for reprobing... if the mild is not effective. You may choose to make your own stripping solution using a recipe like ... Assess the stripping Now it’s time to check to make sure the antibodies from your first round of detection...

...biological. Even the most high-fidelity polymerases can make mistakes and lead to the accumulation of errors ...ensure the entire team is using the same rubric and making the same decisions when analyzing the wide variety...the QC team leaves a note on the plasmid page to make requesting scientists aware of the identified discrepancy...

...Zetsche B, Gootenberg JS, Abudayyeh OO, Slaymaker IM, Makarova KS, Essletzbichler P, Volz SE, Joung J,...plasmids from this publication at Addgene. 4. Makarova KS, Wolf YI, Alkhnbashi OS, Costa F, Shah SA, ...

...meeting in person is often much more productive and makes it easier to build relationships. Nonetheless, the...responsibilities Being a leader in an organization, you need to make sure the organization speaks in “one voice” whilst...such as leadership skills can only assist you and make you more qualified for future career opportunities...

... who keep changing their minds and have trouble making decisions. In my work as a thesis coach for graduate...be reviewed, bring them to the meeting. This will make it easy for your supervisor to support you. 3) Explain...articles on our blog Read Joanne Kamen's Post on What Makes a Good Mentor Resources on the Addgene Wesbite ...

...School of Medicine. In the Lindberg Lab we often make cell lines that overexpress genes of interest; more...Parents of infants will have no problem with this!) Make sure your colonies are well separated. For this,...taking both early- and late-arising clones, and making sure we always maintain a backup replicates of ...

...system), and the third post will describe how you can make your own mutant flies. Find Drosophila Resources...bristly hairs. Females do not have sex combs. To make genetic crosses properly, it is important to isolate...emerging at night. An incubator with a light cycle makes for very predictable work! When flies first emerge...

...the localization of your protein of interest is to make it detectable. Visualization can be achieved using... and their potential overlap, it is essential to make sure that the selected FPs do not influence each...your next localization experiment and invite you to make any fluorescent protein constructs you create in...

...usurped old methods or complemented them in ways that make them much easier. A special class of FPs, the FP...functionality with a single gene Although it is possible to make a timer system composed of two separate FPs (one... This one protein method simplifies cloning and makes your experimental setup less cumbersome. 3. Portability...

...Knockout/Knock-in plasmids Gene targeting methods also make it possible to insert, or knock-in, any gene, tag...instead want to create a conditional knockout. To make a conditional knockout, researchers often use the...floxed alleles. Although CRISPR is very good at making knockouts, knocking in large sections of DNA can...

...m going to share what I’ve learned and hopefully make your decision a little easier if an industry postdoc... your goal is to enter academia, it is easier to make that transition from an academic setting—especially...company you choose. If publishing is important to you, make sure that your project will not involve proprietary...

...capacity and stable expression of lentiviral vectors make them an attractive option. However, safety requirements...has psuedotypes. Psuedotyping is the process of making a hybrid AAV, with the genome from one serotype...Klingler, E., Kuhn, M., Farouni, R., Lagler, S., Kalamakis, G., Lischetti, U., Jabaudon, D., & Platt, R. ...

... any features of the library backbone that would make the library more difficult to work with. For example...bacteria can grow and compete in the same container, making it easier for a smaller, recombined plasmid to ... reference protocols There are many decisions to make when developing your amplification protocol! We ...