Plastid Transformation via Intron-Derived RNA Intermediaries
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Solution Overview
Problem
Existing plastid transformation methods in plants are inefficient due to low transformation efficiency and patchy delivery of genetic information, primarily because they do not utilize endogenous cellular processes for RNA transfer into the plastid genome.
Innovation Solution
Adapting endogenous cellular processes to transfer polynucleotides from the cytoplasm into plastids by inserting polynucleotide sequences into introns, which are then expressed as RNA, reverse transcribed, and integrated into the plastid genome using a multifunctional protein and a reverse transcriptase, facilitating efficient expression of exogenous proteins.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If prior art plastid transformation methods are used, then transformation can be achieved, but transformation efficiency is low and protein production per cell is low
Solution Approach 1:
The patent uses an intron-derived RNA vehicle as an intermediary to transfer genetic information into the plastid. The RNA is transcribed from a nuclear gene, processed through endogenous cellular pathways, and delivered to the plastid where it is reverse transcribed and integrated into the plastid genome. This intermediary approach significantly improves transformation efficiency compared to direct DNA transformation methods.
Solution Approach 2:
The system utilizes endogenous cellular processes for RNA processing, transport, and delivery to the plastid. The cell's own machinery transcribes the nuclear gene, processes the RNA, and delivers it to the plastid, eliminating the need for complex external delivery systems and improving both efficiency and consistency.
2Reliability
If prior art transformation methods are used, then plastid transformation is achieved, but delivery of genetic information is patchy and inefficient
Solution Approach 1:
The intron-derived RNA acts as a reliable intermediary that ensures consistent delivery of genetic information to the plastid. The RNA vehicle is processed through endogenous pathways that naturally ensure uniform distribution and integration, eliminating the patchy delivery problem of prior methods.
Solution Approach 2:
The patent changes the state of genetic material from direct DNA transformation to RNA-mediated transformation. This parameter change from DNA to RNA allows utilization of endogenous RNA processing and transport mechanisms, resulting in more consistent and efficient delivery to the plastid genome.
3Productivity
If endogenous cellular processes are utilized for RNA transfer, then transformation efficiency is enhanced, but process complexity increases
Solution Approach 1:
The system leverages the cell's existing endogenous machinery for RNA transcription, processing, and transport to the plastid. Rather than introducing complex external delivery systems, the invention uses the cell's own self-service capabilities, thereby improving efficiency without significantly increasing overall process complexity.
Solution Approach 2:
The intron-derived RNA vehicle can be used across different plant species and plastid types, utilizing universal endogenous cellular processes. This multi-functional approach allows the same basic mechanism to work in various contexts, simplifying the overall process while maintaining high transformation efficiency.
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 method significantly enhances the transformation efficiency of plastids and ensures consistent expression of exogenous proteins by leveraging natural cellular pathways, leading to improved production of heterologous proteins in plants.
Implementation Method 1
the RNA sequence is reverse transcribed into DNA which then inserts into the plastid genome
Data Source
AI summary
Method for heterologous protein production in plant cell plastids comprising introducing into plant cells nucleic acid components that encode heterologous proteins under the control of promoters operative in plastids, vectors, host cells, plants and uses thereof.