gRNA Mutant Narrowing Base Editor Editing Window
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Solution Overview
Problem
Current CRISPR/Cas systems face challenges in achieving precise single-base editing due to large editing windows, leading to random insertions or deletions, which limits their application in correcting pathogenic point mutations.
Innovation Solution
A method for constructing a gRNA mutant by mutating the guide sequence region to narrow the editing window of base editors, allowing for precise editing of one base by screening for specific mutations that reduce the editing window to a single nucleotide.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If conventional CRISPR/Cas systems are used for gene editing, then DNA cleavage and repair can be achieved, but the editing window is large causing random insertions or deletions instead of precise single-base editing
Solution Approach 1:
The patent introduces a mismatch between the guide RNA and target DNA sequence to shift the editing window. By changing the binding parameters of the guide RNA (introducing deliberate mismatches), the system redirects the base editor's activity to a specific single base position, thereby achieving precise single-base editing while avoiding random insertions or deletions throughout the large original editing window
Solution Approach 2:
The patent uses a specially designed guide RNA as an intermediary element that mediates between the base editor and the target DNA. This guide RNA contains deliberate mismatches that act as a positioning mechanism, directing the base editor to edit only the specific base at the mismatch position rather than allowing editing across the entire editing window, thus resolving the precision problem
2Adaptability or versatility
If base editors with large editing windows are used, then multiple bases can be edited, but it is difficult to correct specific pathogenic point mutations without additional base modifications causing side effects
Solution Approach 1:
The patent changes the binding parameters of the guide RNA by introducing deliberate mismatches at specific positions. This parameter change narrows the effective editing window to a single base position, allowing precise correction of pathogenic point mutations without causing unwanted modifications to other bases within the original editing window, thereby eliminating side effects
Data Source
AI summary
Provided are a gRNA mutant and the use thereof, and further provided are a method for constructing same and a base editor containing same. The gRNA mutant is used in the base editor, and can universally reduce a base editing window, thereby improving the specificity of gene editing, and achieving specific editing on one base.