Author: Long T. Nguyen; Brianna M Smith; Piyush K. Jain
Title: Enhancement of trans-cleavage activity of Cas12a with engineered crRNA enables amplified nucleic acid detection Document date: 2020_4_14
ID: n5kpvsvg_5
Snippet: Based on our findings that the trans-cleavage activity is drastically improved by 7-mer ssDNA extensions to the 3'-end of crGFP, we questioned if the binding of crRNA with LbCas12a itself is influenced by such modifications. A biolayer interferometry binding kinetic assay revealed that the dissociation constant, Kd, between the binary complex LbCas12a:crRNA and LbCas12a:crRNA+3'DNA7 are comparable within a low nM scale ( Fig. 2i and Supplementary.....
Document: Based on our findings that the trans-cleavage activity is drastically improved by 7-mer ssDNA extensions to the 3'-end of crGFP, we questioned if the binding of crRNA with LbCas12a itself is influenced by such modifications. A biolayer interferometry binding kinetic assay revealed that the dissociation constant, Kd, between the binary complex LbCas12a:crRNA and LbCas12a:crRNA+3'DNA7 are comparable within a low nM scale ( Fig. 2i and Supplementary Fig. 9 ). These binding results suggest that the 3'DNA7 modification on crRNA does not affect the binary complex formation between the LbCas12a and the crRNA.
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