P-Domain Mutations for Specific Nucleic Acid Hybridization
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Solution Overview
Problem
Existing genome engineering methods face challenges in achieving precise and specific binding of nucleic acid-targeting nucleic acids, leading to non-specific interactions and inefficient modification of target nucleic acids.
Innovation Solution
Engineered nucleic acid-targeting nucleic acids with mutations in the P-domain or bulge region, allowing for improved hybridization and specificity, as well as the use of site-directed polypeptides and donor polynucleotides for targeted modifications.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If wild-type nucleic acid-targeting nucleic acids are used, then the system is simple and easy to manufacture, but binding specificity and precision are insufficient leading to non-specific interactions
Solution Approach 1:
The patent applies parameter changes by introducing specific point mutations in the P-domain of the nucleic acid-targeting nucleic acid. These mutations alter the nucleotide sequence parameters to improve binding specificity and reduce non-specific interactions, directly resolving the contradiction between binding precision and sequence complexity
Solution Approach 2:
The patent applies local quality by making targeted modifications only in the P-domain region of the nucleic acid-targeting nucleic acid, while maintaining the rest of the sequence (including the guide region and tracrRNA portion) as wild-type. This localized approach improves binding specificity without unnecessarily complicating the entire molecule
2Productivity
If wild-type nucleic acid-targeting nucleic acids are used, then the system is simple and easy to operate, but modification efficiency is low due to non-specific binding
Solution Approach 1:
The patent introduces point mutations in the P-domain to enhance binding specificity, which directly improves modification efficiency by reducing non-specific binding events. This parameter change enables the system to achieve higher productivity without significantly complicating operation
Solution Approach 2:
The patent utilizes the transient nature of the nucleic acid-targeting nucleic acid complex, where the engineered mutations ensure that only specific target nucleic acids form stable complexes, while non-specific interactions are minimized. This approach improves efficiency without requiring complex operational protocols
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
Enhances the specificity and efficiency of nucleic acid modifications, reducing non-specific binding and enabling precise genetic manipulations.
Implementation Method 1
the engineered nucleic acid-targeting nucleic acid is adapted to hybridize to a target nucleic acid
Data Source
AI summary
This disclosure provides for compositions and methods for the use of nucleic acid-targeting nucleic acids and complexes thereof. Genome engineering can refer to altering the genome by deleting, inserting, mutating, or substituting specific nucleic acid sequences. The altering can be gene or location specific. Genome engineering can use nucleases to cut a nucleic acid thereby generating a site for the alteration. Engineering of non-genomic nucleic acid is also contemplated.