Algal Methyltransferase Mutation for Exogenous Gene Expression
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Solution Overview
Problem
Algal species face challenges in expressing exogenous genes due to epigenetic mechanisms like DNA methylation, particularly CHG and CHH methylation, which silences foreign DNA, limiting the efficiency of gene expression.
Innovation Solution
Mutating or attenuating methyltransferase genes responsible for CHG and CHH DNA methylation in algal organisms, such as those encoding CHG DNA methyltransferase, to reduce methylation levels and enhance the expression of exogenous genes, using methods like CRISPR/Cas systems for genetic engineering.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If DNA methylation mechanisms are active in algal species, then native genome stability is maintained, but exogenous gene expression is silenced
Solution Approach 1:
The patent applies local quality by differentiating between native and exogenous DNA regions. The methyltransferase system naturally distinguishes between self (native) and non-self (exogenous) DNA, applying methylation selectively to exogenous sequences while preserving native genome stability. This localized action allows simultaneous maintenance of genome integrity and enhancement of transgene expression.
Solution Approach 2:
The patent converts the harmful silencing effect of DNA methylation into a beneficial tool for gene expression control. By understanding and manipulating the methylation mechanism, the invention creates mutant algae with reduced methylation activity that specifically benefit exogenous gene expression while maintaining native genome stability, effectively turning a defensive mechanism into a productive asset.
2Productivity
If methyltransferase genes are mutated or attenuated, then exogenous DNA methylation is reduced and gene expression is enhanced, but potential genomic instability may occur
Solution Approach 1:
The patent applies partial action by creating mutants with attenuated rather than completely eliminated methyltransferase activity. The partial reduction in methylation capability is sufficient to enhance exogenous gene expression while maintaining enough activity to preserve native genome stability, avoiding the risks of complete methylation suppression.
Solution Approach 2:
The patent changes the parameter of methyltransferase activity from normal levels to attenuated levels through mutation. This parameter change reduces CHG and CHH methylation specifically affecting exogenous DNA while preserving sufficient activity for maintaining native genome integrity, achieving a balance between enhanced transgene expression and genomic stability.
3Manufacturing precision
If CRISPR/Cas systems are used for gene editing, then precision in mutating methyltransferase genes is improved, but system complexity increases
Solution Approach 1:
The patent applies self-service by utilizing the CRISPR/Cas system's inherent ability to perform precise gene editing without requiring additional complex intervention mechanisms. The system uses guide RNA to automatically target and edit the methyltransferase genes with high precision, reducing the need for complex external control systems and simplifying the overall genetic engineering process.
Data Source
AI summary
The present application relates to the identification of novel DNA methyltransferases including CHG methylation in algal species. The present application relates to algal mutants permitting the expression of exogenous genes by alleviating the epigenetic mechanisms of CHG and CHH methylation of exogenous DNA and mono- and tri-methylation of lysine 9 of histone 3 (H3K9). This is achieved by mutating or attenuating the methyltransferase (MTase) genes in algae. The present application also relates to methods for efficiently expressing exogenous genes in algal species.