Universal Promoter Sequences for Multi-Species Enzyme Expression
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current methods for evaluating and expressing enzymes across multiple host species are hindered by the need for species-specific promoters, leading to high costs and inefficiencies, especially when dealing with unknown gene sequences or toxic enzymes, and lack a universal promoter system that functions across a wide range of industrially relevant species.
Innovation Solution
Development of polynucleotide sequences with specific promoter activity that can drive expression in a wide range of both prokaryotic and eukaryotic species, allowing for the use of a single expression cassette to test enzyme activity across multiple hosts and enabling efficient selection marker systems for both forward and backward selection.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If species-specific promoters are used for each host, then expression efficiency in individual species is improved, but device complexity and cloning workload increase significantly when testing across multiple species
Solution Approach 1:
The patent develops a universal promoter system that functions across multiple species (E. coli, S. cerevisiae, P. chrysogenum, B. subtilis) using a single promoter sequence. This multi-functional promoter eliminates the need for species-specific promoters, thereby reducing cloning workload while maintaining expression efficiency across diverse hosts.
Solution Approach 2:
The invention merges the functionality of multiple species-specific promoters into a single universal promoter construct. By combining the essential regulatory elements needed for transcription initiation across different species into one promoter sequence, the system achieves broad applicability without requiring separate expression cassettes for each host.
2Reliability
If synthetic DNA is used for codon optimization, then gene expression in new host species is improved, but manufacturing costs increase to around 1000 US dollars per gene
Solution Approach 1:
The universal promoter system enables direct testing of native or near-native gene sequences across multiple species without requiring expensive synthetic codon-optimized versions. By using a promoter that functions in diverse hosts, the system achieves successful expression with minimal sequence modification, dramatically reducing DNA synthesis costs.
Solution Approach 2:
The invention employs a cost-effective approach by using simple, modular promoter sequences that can be readily synthesized and tested. Rather than investing in expensive custom-designed codon-optimized genes, the system uses affordable promoter constructs that can be rapidly deployed across multiple species to evaluate enzyme function.
3Productivity
If Gateway recombination systems are used to transfer genes, then gene transfer efficiency is improved, but nucleotide insertions (20-30 bp) severely hamper transcription and translation
Solution Approach 1:
The patent extracts and eliminates the problematic 20-30 bp nucleotide insertions introduced by Gateway recombination systems from the expression cassette. By removing these interfering sequences while retaining the essential gene and promoter components, the system achieves both efficient gene transfer and optimal transcription/translation without the hampering effects of extraneous nucleotides.
Data Source
AI summary
The present invention relates to polynucleotide sequences which enable a polynucleotide control sequence, such as a promoter, to direct expression in a wide range of industrially relevant species, both prokaryotes and eukaryotes. When the polynucleotide sequences of the invention are applied in combination with selection marker genes it is possible to perform selectable cloning in a laboratory host and use the same construct in the final host.