...non-dividing cells. The first lentiviral vector system consisted of three plasmids: the packaging, envelope, and...capacity of a lentiviral vector but for which transient transgene expression is sufficient. Lentiviral...

...expression Whether BiFC constructs are expressed transiently by transfection or stably impacts the success...independent of PPIs. Simply compare the fluorescence intensity of your BiFC constructs to your negative control...

...enzymes do not contain the RuvC and HNH domains responsible for DNA cleavage, so they cannot directly edit.... PspCas13b does not require a PFS, and it is sensitive to mismatches in target RNA from bases 12-26 of...

...biologist--as an example of “big science,” a massive, expensive project that required collaboration from scientists...five Addgene colleagues who read it with me, the consistent response from scientist and non-scientist alike...

...For these reasons, we are in the process of transitioning our titering method from qPCR to ddPCR. Droplet...titer your AAV by ddPCR. Droplet digital PCR is a sensitive assay, so cross contamination into a no template...

...category, will leave us with a single plasmid to consider. If you get this far and realize you need to apply...and let us help.  Check back on Thursday for an inside look at our mascot and homepage redesign. Additional...

...that is wide enough to dunk your plant’s flowers inside. 50 mL falcon tubes or plastic weigh boats are ...from a colony on a plate will reach an optimal density in 18-24 hours. I then add 0.5 mL of this starter...

...human CRISPRko sgRNA library The Brunello library consists of 77,441 sgRNAs with an average of 4 sgRNAs per...Mediator) uses an improved activator design that consists of inactive Cas9-VP64, a sgRNA with two MS2 RNA...

...Saccharomyces cerevisiae in 1989 and are still used extensively to screen for molecular interactions in the cell...post-translationally modified, co-expression of the enzyme responsible for the modification in the assay host strain...

... Although we are busy with many other work responsibilities, we make team building activities a priority...at Addgene. In addition to her operational responsibilities, she enjoys organizing team building events...

...deposited by David Root of the Broad Institute) has consistently been one of the repository’s most frequently...Rory Macneil on Bitesize Bio Data Freedom: The Expansion of Data Sharing in Research Publications by Jim...

... through a 0.22 μm membrane. Store at 4 °C. Considerations Before You Start The health of the HEK293T ...vigorously multiple times to obtain a single cell suspension (no clumps of cells). Pool cells from 2 x T-175...forth or shake vigorously to obtain a single cell suspension (no clumps). Count cells using a hemocytometer...above. Add 50 units of benzonase per mL of viral suspension (you should have ~18 mL from steps 12 & 13). ...

...our own repository data, giving us significant insight into which vectors are likely to be of most use...Swanson, L. E., & Fan, M. (2016). Practical Considerations for Using Pooled Lentiviral CRISPR Libraries...

...plasmids tend to be high copy number. Here, we will consider the replication mechanism of plasmids under stringent...plasmid to balance the total plasmid number.  Let’s consider pSC101. This plasmid is stringently controlled...

...have, then bring them to your mentor. Pro tip! Consider physically walking through the protocol, by going...receive help until you’ve already started a time sensitive experiment. Ready to fly solo? Let your bench...

...from many other types of cancer. Mesothelioma is considered an orphan cancer; it is rare disease with only...non-resectable mesothelioma by the National Comprehensive Cancer Network, a coalition of the top 27 cancer...

...gain insight into protein structure and function (Pan et al., 2018). cDNA libraries, which consist of plasmids...

...d like to manipulate in mind. Now it’s time to consider where you would like to attack this gene. Should...sequences into HEK293T cells. The fluorescence intensity was classified into four groups (4; brighter than...

...editing field by making experimental workflows considerably easier, faster, and more efficient than previous...novice to expert-level researchers in achieving consistently successful CRISPR experiments. Jessica is a ...

...CRISPR/Cas Toolkit for plants includes 95 plasmids consisting of CRISPR/Cas nucleases, base editors, gRNA backbones...activation-based norepinephrine (GRABNE) sensors allow for sensitive and high spatiotemporal measurements of norepinephrine...