Gene Expression System with Splice Control Sequences
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Solution Overview
Problem
Current gene expression systems lack the ability to achieve precise, tissue-specific, stage-specific, and sex-specific control of protein expression, which is essential for applications such as pest control and therapeutic interventions, due to the limitations of existing promoters and splicing mechanisms.
Innovation Solution
A gene expression system that combines a coding sequence with a promoter and splice control sequences, specifically intronic sequences, to mediate alternative splicing, allowing for sex-specific, stage-specific, and tissue-specific expression of proteins, using mechanisms like the Cctra intron system, which enables the differential expression of proteins by controlling alternative splicing patterns.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If conventional promoters are used to control gene expression, then expression can be achieved, but precise tissue-specific, stage-specific, and sex-specific control cannot be obtained
Solution Approach 1:
The invention segments the gene expression control into multiple independent functional modules: a promoter region for basal control, intronic sequences containing splice regulatory elements for alternative splicing control, and specific splicing factors. This segmentation allows each module to be independently optimized and combined to achieve precise tissue-specific, stage-specific, and sex-specific expression patterns that conventional single-promoter systems cannot provide.
Solution Approach 2:
The invention adds a new dimension of control by incorporating alternative splicing mechanisms alongside traditional promoter control. This creates a multi-layered regulation system where the same gene can produce different protein isoforms through alternative splicing in different tissues and developmental stages, dramatically increasing the versatility and precision of gene expression control without requiring multiple separate promoter systems.
2Manufacturing precision
If alternative splicing mechanisms are introduced to achieve specific expression patterns, then expression control is improved, but system complexity increases
Solution Approach 1:
The invention utilizes universal splicing regulatory elements and conserved splicing factor mechanisms that can be applied across different genes and expression contexts. The intronic sequences contain universal splice site recognition motifs and regulatory elements that work with the cell's existing spliceosome machinery, allowing precise expression control without requiring entirely new molecular components or drastically increasing system complexity.
3Manufacturing precision
If multiple control mechanisms are combined for precise expression, then expression specificity is improved, but the number of components increases
Solution Approach 1:
The invention merges multiple control functions into a single integrated gene construct. The promoter, intronic sequences with splice regulatory elements, and coding region are combined into one continuous genetic unit that functions as a unified system. This merging achieves high expression specificity through coordinated action of the integrated components while avoiding the need for separate independent control systems that would increase the total number of genetic components.
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 system allows for the precise and controlled expression of proteins in specific contexts, such as female-specific expression in mosquitoes, enabling applications like anti-filarial effector molecule expression in adult female mosquitoes, thereby addressing the limitations of existing systems in achieving desired expression patterns.
Implementation Method 1
Alternative splicing is also known as pre-mRNA splicing and involves the removal of one or more introns and ligation of the flanking exons. This reaction is catalyzed by the spliceosome, a macromolecular machine composed of five RNAs and hundreds of proteins
Data Source
AI summary
A gene expression system is provided. The system comprises at least one coding sequence to be expressed in an organism, and at least one promoter operably linked thereto. It further comprises at least one splice control sequence which, in cooperation with a spliceosome, mediates alternative splicing of RNA transcripts of the coding sequence. The mediation of alternative splicing is in a sex-specific, stage-specific, germline-specific and tissue-specific manner.