...worthwhile. Particularly if in the end all they want to do is to quickly generate a single cell line and proceed... Advisory Board) says: "When you're setting up to do a genetic screen, you need to focus on building a...assay is scalable and robust. What you don't need to do is spend your time troubleshooting reagent production...

...enable efficient base editing and prime editing. How do you then set up an MBE system that contains a double-win...editing, while keeping the system to minimal size. To do so, they harnessed the tRNA processing mechanism,...exhibited reduced Cas9-dependent off-target editing and do not cause higher Cas9-independent off-target editing...

...journey can be as exciting as it is challenging. How do you prepare for this shift and navigate the pitfalls...those accustomed to hands-on work. The temptation to "do it yourself" can result in burnout and underutilized...managerial roles, and I've witnessed several colleagues do this while changing industries and positions. Some...

...vector propagation and sequencing a headache. How do I maintain ITR integrity and prevent unwanted mutations...reduce the likelihood of ITR deletions occurring.  How do I validate my ITR sequences?  Next-generation sequencing...immediately outside of the ITRs in the plasmid backbone and do not properly assemble, but these sequences can sometimes...

...in the article are those of the guest blogger and do not necessarily represent the views of Addgene. I... 3. NEB Tools by New England BioLabs In my lab we do a lot of genetic construction and cloning experiments...learn about biochemistry. The least this app will do is allow you to write your colleagues’ names in a...

...introduces himself and explains why he decided to do an MD PhD - it's all in the family. 3:56 - 7:04 :...gene drives work and what work he's planning on doing with CRISPR-based gene drives focusing on safety...

...Duedu inspire new Ghanaian scientists to continue to do excellent work in the biological sciences. Future...plasmids from the Addgene community and hope we can do even more to enable scientists in developing regions...

...they do in the software. Most sgRNA design tools require the user to specify an organism. They do this...what the parameters represent. The best learn-by-doing tool is E-CRISP. The authors have made a huge effort...for labs: Define what you need the software tool to do before going shopping for one. Use the CRISPR Software...criteria. Repeat until you have found the best tool. Do several experiments and use the results to re-evaluate...make a difference in our experiments and lives. To do this effectively, there needs to be good communication...

... cost. The problem with these assays is that they do not provide sequence-level data. They also have a... a limit of detection of ~5%. This means they do not reliably detect editing events that occur in less...variable) Sequence-level data, extremely sensitive Do not sequence all DSBs, may miss unpredicted off-target...Tsai et al. 2015). Additionally, current algorithms do not take into account other elements, including those...GUIDE-seq and BLESS detect double-stranded breaks and do not require high sequencing read counts making them...

... was doing the kind of work my lab wanted to do already seemed to know each other and what to do.  Saved...tracks), something we didn’t have enough people to do in the pilot. We also found that all participants...

...take to reduce them. But we do know we will continue to ask “What can we do to make our materials both ...by researchers all over the world — and we really do mean all over (Figure 3). We continue to add new ...more reproducible science and learn what Addgene can do to help. More resources on the Addgene blog Twenty...

...video background! (720 px or 1080 px).         Do you do a lot of transformations with the plasmids you...

...previously identified Cas13a-c orthologs, but they do include HEPN nuclease domains characteristic of the...here Find the Konermann et al. Plasmids here How do Cas13d enzymes compare to other Cas13s? Like other... and is thus HEPN-independent. These enzymes also do not require a protospacer flanking sequence, so you...

...need to sort through them to find the desired clone. Do your selection, then screen Why add a screen to your...plasmids that contain the gRNA (white) from those that do not (blue). Let’s take a look at a few screening ... containing plasmids with inserts from those that do not is by using a restriction digest. The key is ...

...Golden Gate etc.), and try to find the simplest way to do it (i.e. avoid assembling too many fragments at once...by methylation like XbaI, ClaI, etc. If you are doing multiple digestion be sure that the buffers and ...plasmid, and analyze it using the following steps: Do a single or double digest to be sure that the plasmid...

...groups and individuals with similar ideas to actually do something about it. Building a diverse team with ...There are many brilliant minds out there, but how do you get them to connect? Through networking of course...solving a community’s issue without involving them can do more harm than good. Check out this webinar for more...

...we understand what genes do! Viruses have several options to allow you to do exactly that: shRNAs are...clinical promise moving forward. What viruses can do for you Whether you are interested in basic science...long-lasting immune response even after a single dose. Vaccine development strategies. Image courtesy...

...inducible? How long do you need control? Will temporary control (a few days) be enough, or do you want a permanent...101: Repressible Promoters CRISPR 101: Which Cas9 Do I Choose for my Experiment Additional resources ...

...phenotype in the surviving population. Selections do require there to be a culture condition that can ...sound like a great shortcut, right? Who would ever do a screen when a selection can get you to the answer...culture condition which eliminates manipulations that do not affect the function of interest. An example where...culture cells in the drug of interest at a lethal dose for non-manipulated cells and then assay cells which...

...” of mentors as one key to their success. But how do you find these elusive teachers, supporters and advisors...be read or watched during the meeting.  Scientists do better with process, so don’t just get together and...blog in this series will have more ideas on what to do once you are in a mentoring relationship.  Celebrate...