Codon-optimized Cry1Da Gene for Resistant Pest Control
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Solution Overview
Problem
Current transgenic plants expressing Cry proteins, such as those with the cry1F gene, are no longer effective in controlling pest species like Spodoptera frugiperda and Diatrea saccharalis due to resistance development, necessitating a new strategy for efficient pest control.
Innovation Solution
Development of codon-optimized cry1 Da nucleic acid molecules with optimized G+C content, integrated into transgenic maize plants using Agrobacterium-mediated transformation, which enhances the expression and efficacy of the Cry1 Da protein against resistant pest populations.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If native cry genes from B. thuringiensis are inserted into plants, then insecticidal activity against lepidopteran pests is achieved, but gene expression levels are too low to provide adequate protection
Solution Approach 1:
The patent applies codon optimization to change the nucleotide sequence parameters of the cry1Da gene while maintaining the same amino acid sequence. This involves substituting codons to match plant preference patterns, adjusting GC content to 50-60%, and modifying codon usage frequency to align with maize codon bias, thereby enhancing translation efficiency and protein expression levels without altering the insecticidal function
Solution Approach 2:
The patent introduces plant-specific regulatory elements at specific locations in the gene construct, including the maize ubiquitin promoter (Ubi1) for transcriptional control and the nopaline synthase terminator (NOS) for proper mRNA processing. These localized modifications optimize gene expression in the plant system while preserving the cry gene's core insecticidal functionality
2Reliability
If transgenic plants with existing cry genes are used, then pest control is provided, but resistance development in pest populations occurs reducing effectiveness
Solution Approach 1:
Instead of using the commonly deployed cry1F gene that pests have developed resistance against, the patent inverts the approach by selecting and optimizing the cry1Da gene variant. This alternative cry gene provides a different mode of action that is effective against resistant pest populations, essentially reversing the problem-solution dynamic by switching from a failed approach to a successful alternative
Solution Approach 2:
The patent creates a composite gene construct that integrates the optimized cry1Da coding sequence with multiple plant-compatible regulatory elements (ubiquitin promoter, NOS terminator, and other enhancers). This composite structure ensures high-level expression of the insecticidal protein while maintaining stability and effectiveness against resistant pests over extended periods
3Productivity
If B. thuringiensis cry genes are expressed in plants, then insecticidal protein is produced, but codon incompatibility between bacteria and plants reduces expression efficiency
Solution Approach 1:
The patent systematically changes the nucleotide sequence parameters of the bacterial cry1Da gene to match plant characteristics. This includes adjusting codon usage frequency to reflect maize codon bias, modifying GC content to fall within the optimal 50-60% range for plant expression, and ensuring proper codon-anticodon pairing efficiency in the plant cellular environment, thereby dramatically improving protein production levels
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
The codon-optimized cry1 Da gene construct achieves high expression levels in maize plants, providing effective control over Spodoptera frugiperda and Diatrea saccharalis, including resistant populations, thereby overcoming the limitations of existing transgenic crops.
Implementation Method 1
integrated into transgenic maize plants using Agrobacterium-mediated transformation
Data Source
AI summary
The present invention relates to new codon-optimized cry1Da nucleic acid molecules from a gene sequence isolated from bacterium Bacillus thuringiensis. These molecules are used in the preparation of nucleic acid constructs, vectors and host cells, allowing the production of transgenic plants, such as corn, resistant to invertebrate pests, such as insects from the order Lepidoptera, particularly Spodoptera frugiperda (Noctuidae, Lepidoptera) and Diatrea saccharalis (Crambidae, Lepidoptera). Plant cells and transgenic plants comprising the molecules or constructs of the invention are also objects of the present invention. In particular, the transgenic plants according to the present invention are able to control caterpillars of the cited species that have become resistant to plants containing the cry1F gene. In addition, the present invention relates to a method for transforming a cell, a method of controlling invertebrate pests in crop plants and uses of nucleic acid molecules or constructs in the production of transgenic plants and for controlling invertebrate pests.