...individually or simultaneously. Do note for control purposes that there are dosage effects. For example, overexpression... of interest, the next question to ask is what to do with them. In some cases, there might be a mutation...will need to be overexpressed or knocked down. To do this, Drosophila geneticists use the Gal4/UAS system...

...approaches everything; we asked ourselves, “How can we do this in a way that most helps scientists and allows... the service and figure out what we can afford to do and when. Addgene’s Legal Team acquired permission...impression that it took a lot of work and dedication to do this well ... because it did. So finally, a big thank...

...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...

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

...protein present on the plasmid. gRNA plasmids that do not co-express a Cas protein require a separate plasmid...multiple target genes in Escherichia coli. Koffas Do you have suggestions for other plasmids that should...pyogenes EGFP Zoldos pdCas9-DNMT3A-PuroR 71667 Mammalian U6 yes, methylation S. pyogenes Puro Zoldos pX330-U6... Mammalian U6 yes, methylation S. pyogenes Puro Zoldos pBLO1811_Cas9_noNLS_human 74490 Mammalian U6 yes...

...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 ...

...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 ...

...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...

...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...

...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...

...” 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...

...Unfortunately, many science graduate programs are still doing an inadequate job of supporting science trainee ...into diverse, successful careers.  We’re not even doing a very good job of providing skills training for...

...opportunities to fall in your lap. Sometimes, in order to do the thing you love, you have to create opportunities...honest), I would tell them to follow your passion. Do what you love. Don't worry about the rest.”- Jason...

...maybe smaller Cas systems could exist in nature. To do this, they mined a database of microbial genome and...indiscriminately cleave ssDNA in trans like Cas13 and Cas12 do for RNA and dsDNA, respectively. However the authors...

...Addgene, so we need to be cost and time efficient. We do this by making our own competent cells and using ...each new batch of competent cells, don’t forget to do a negative control (plate your cells without adding...