Can I read Optimized protocols for the characterization of Cas12a activities on EtoBox?
Optimized protocols for the characterization of Cas12a activities by Lindsie Martin; Saadi Rostami; Rakhi Rajan is a book available to read on EtoBox.
What is Optimized protocols for the characterization of Cas12a activities about?
by a 20 nucleotide (nt) segment that is complementary to the "guide" region of its CRISPR RNA (crRNA). The guide sequence of the crRNA can be programmed to target any DNA with a cognate PAM and is the basis for Cas12a's current use for gene editing in numerous organisms and for medical diagnostics. While Cas9 (type II effector protein) is widely used for gene editing, Cas12a possesses favorable features such as its smaller size and creation of staggered double-stranded DNA ends after cleavage that enhances cellular recombination events. Collected here are protocols for the recombinant purification of Cas12a and the transcription of its corresponding programmable crRNA that are used in a variety of Cas12a-specific in vitro activity assays such as the cis, the trans and the guide-RNA independent DNA cleavage activities with multiple substrates. Correspondingly, protocols are included for the quantification of the activity assay data using ImageJ and the use of MATLAB for rate constant calculations. These procedures can be used for further structural and mechanistic studies of Cas12a orthologs and other Cas proteins.
- Author
- Lindsie Martin; Saadi Rostami; Rakhi Rajan
- Publisher
- Academic Press, an imprint of Elsevier
- Published
- 2023
- Language
- EN
- ISBN
- 9780323992657
- Subjects
- Science, Chemistry, Biology