In vitro Cas9 RNP assembly and genomic DNA extraction
This protocol is extracted from research article:
Universal intracellular biomolecule delivery with precise dosage control
Sci Adv, Oct 31, 2018; DOI: 10.1126/sciadv.aat8131

The Cas9 RNP was prepared according the protocols described previously (42). The Cas9 RNP was made immediately before the experiment. To make the Cas9 RNP, purified Cas9 protein was incubated with sgRNA at a 1:1.2 molar ratio in 20 μM Hepes at pH 7.5, 150 mM KCl, 1 mM MgCl2, 10% glycerol, and 1 mM TCEP at 37°C for 10 min. Cells were incubated at 37°C for 24 to 48 hours after Cas9 RNP delivery. To extract the genomic DNA, the cells were lysed by 20 to 100 μl of QuickExtract solution (Epicentre) at 65°C for 20 min and then at 95°C for 20 min and were stored at −20°C. The concentration of genomic DNA was measured by NanoDrop (Thermo Scientific).

Note: The content above has been extracted from a research article, so it may not display correctly.



Q&A
Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.



We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.