Ribozyme Scaffold RNA Cleaving for Plant DNA Demethylation
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Solution Overview
Problem
Current methods for directing demethylation of target DNA in plants are inconvenient and not rapid, lacking specific control over methylation in DNA sequences, which is essential for manipulating gene expression and creating plants with desired traits.
Innovation Solution
The method involves cleaving scaffold RNA produced by transcription of target DNA using specific ribozymes in plant cells, thereby inhibiting methylation of target DNA in plants, allowing for specific control of gene expression and trait manipulation without relying on recombinant technology.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If dCAS-fused TET1 is used to direct demethylation, then specific demethylation of target DNA is achieved, but the process becomes complex and not rapid due to requiring recombinant technology
Solution Approach 1:
The invention uses scaffold RNA as an intermediary to guide the ribozyme to the target DNA sequence. The ribozyme cleaves the scaffold RNA, which indirectly achieves demethylation of the target DNA without requiring direct interaction between the demethylating enzyme and the DNA, thus simplifying the system while maintaining specificity
Solution Approach 2:
The invention replaces the complex recombinant dCAS-TET1 system with a ribozyme-based system that uses RNA cleavage to achieve the same demethylation effect. This substitution of mechanism (from direct enzymatic demethylation to RNA cleavage-mediated demethylation) simplifies the overall process while maintaining target specificity
2Productivity
If epigenetic control is manipulated to increase accumulation of functional components, then productivity is improved, but the complexity of controlling methylation increases
Solution Approach 1:
The invention applies demethylation locally to specific target DNA sequences rather than globally affecting the entire genome. The ribozyme is designed with specific sequence recognition capability that targets only the desired loci, allowing precise local modification of epigenetic states to enhance functional component accumulation without complex global control
Solution Approach 2:
The ribozyme system is self-regulating through the availability of scaffold RNA. When the target DNA is transcribed, scaffold RNA is produced and automatically cleaved by the ribozyme, creating a self-amplifying demethylation effect at the target locus without requiring complex external control mechanisms
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
This approach enables convenient and rapid inhibition of methylation, facilitating the production of plants with desired traits and increased accumulation of functional components, thus enhancing the production of useful proteins and metabolic compounds.
Implementation Method 1
cleavage of the scaffold RNA by expression of one or more ribozymes specific for the scaffold RNA in the plant cells
Data Source
AI summary
Plants are created with desired traits by conveniently and rapidly inhibiting methylation of target DNA in plants, without using recombination technology. Scaffold RNA produced by transcription of target DNA is cleaved in an RNA-directed DNA methylation mechanism.