Modified gRNA 5' Cap and PolyA Tail for CRISPR Stability

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Solution Overview

Problem

The CRISPR/Cas system for genome editing in eukaryotic cells faces inefficiencies in gene editing, particularly in certain cell types, due to the stability and functionality of guide RNA (gRNA) components.

Innovation Solution

Modification of gRNA molecules by incorporating a 5' cap structure or a 3' polyA tail, or both, enhances their stability and efficiency in gene editing, with specific chemical modifications and lengths of the polyA tail optimizing performance.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If standard gRNA molecules are used for CRISPR/Cas genome editing in eukaryotic cells, then the system can function to target and edit genes, but the efficiency and stability of gene editing is insufficient, particularly in circulating cells like T cells

Engineering Contradiction:

Improvegene editing efficiencyVSAvoidgRNA stability

Core Design Contradiction:

ProductivityVSReliability

Solution Approach 1:

The patent modifies the chemical structure of gRNA molecules by adding a 5' cap structure (m7GpppG) and/or a 3' polyA tail with specific lengths (5-100 nucleotides). These parameter changes in the molecular structure significantly improve the stability and editing efficiency of gRNA in eukaryotic cells, particularly in circulating cells like T cells, by protecting against degradation and enhancing cellular uptake and function

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The patent creates composite gRNA structures by combining standard gRNA sequences with eukaryotic mRNA protective elements (5' cap and 3' polyA tail). This composite approach integrates features from different molecular systems to produce a gRNA that functions effectively in eukaryotic cells while maintaining the CRISPR/Cas targeting capability

Inventive Principle:

Principle #40Composite materials

2Reliability

If gRNA molecules are modified with 5' cap structures and 3' polyA tails, then stability and editing efficiency are improved, but the molecular complexity and synthesis difficulty increase

Engineering Contradiction:

ImprovegRNA stabilityVSAvoidgRNA molecular structure

Core Design Contradiction:

ReliabilityVSDevice complexity

Solution Approach 1:

The patent incorporates the 5' cap structure and 3' polyA tail into the gRNA molecule design from the outset, rather than adding them after synthesis. This preliminary action during the design and synthesis phase streamlines the production process and ensures the protective elements are properly integrated, reducing overall complexity despite the enhanced molecular structure

Inventive Principle:

Principle #10Preliminary action

Data Source

PatentUS20230002760A1Crispr/CAS-related methods, compositions and components

Publication Date: 2023.01.05 EDITAS MEDICINE INC

AI summary

CRISPR/Cas-related compositions and methods which provide for efficient gene editing of eukaryotic cells using modified gRNAs.