Asymmetric RNAi Library Construction via Non-Palindromic Sequences
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
There is a long-standing need for methods to construct random, inhibitory hairpin RNA libraries, which are difficult to synthesize due to limitations in conventional procedures, making existing libraries fully palindromic and unsuitable for RNAi applications.
Innovation Solution
The development of recombinant expression vectors that express RNA molecules with a double-stranded region of random sequence, including a first region of substantially random sequence, a second non-self-complementary region, and a third region substantially complementary to the first region, allowing for the generation of libraries with partial double-stranded secondary structure.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of manufacture
If conventional procedures for randomization and generation of RNA libraries are used, then the library can be constructed, but the resulting libraries are fully palindromic and unsuitable for RNAi applications
Solution Approach 1:
The patent introduces asymmetry into the RNA library construction by using non-palindromic random sequences. Instead of generating fully palindromic sequences through conventional randomization, the invention designs sequences where the 5' and 3' ends are not mirror images, creating asymmetric structures that are suitable for RNAi while maintaining ease of construction through modified synthesis protocols
2Adaptability or versatility
If fully palindromic sequences are generated through conventional randomization, then the library construction is simplified, but the sequences become unsuitable for RNAi applications
Solution Approach 1:
The patent applies local quality by creating specific regions with different properties within the RNA sequence. The invention uses a modular approach where certain regions are randomized while maintaining specific structural constraints in other regions, allowing the library to have both diversity and functional suitability for RNAi applications
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
Enables the identification of RNA therapeutic molecules and drug targets by generating libraries that are not fully palindromic, making them suitable for RNAi applications and overcoming the limitations of conventional synthesis methods.
Implementation Method 1
obtaining a nucleic acid intermediate II from a single-stranded nucleic acid intermediate I
Implementation Method 2
obtaining a double-stranded intermediate III from the nucleic acid intermediate II, comprising the nucleic acid intermediate II and a nucleic acid molecule that hybridizes with the nucleic acid intermediate II
Data Source
AI summary
This invention provides expression vectors for a ribonucleic acid (RNA) molecule comprising a double-stranded region of random sequence, sets and libraries of same, methods of generating same, and methods for identifying an RNA therapeutic or RNA molecule that has an ability to affect a biological parameter, for identifying a drug target for a disease or disorder of interest, and for identifying a variant of an RNA molecule that has an altered ability to affect a biological parameter of interest.