Monascus Promoter Sequences for High Protein Expression
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Solution Overview
Problem
Current technologies lack effective promoters for high expression of specific genes like phosphoenolpyruvate carboxykinase (acuF) and heat shock protein (Hsp) in Monascus species, limiting their application in protein over-expression and recombinant DNA technology.
Innovation Solution
Identification and characterization of strong promoter regions upstream of the acuF and Hsp genes, which are then used to create recombinant DNA molecules and expression vectors for enhanced protein expression in Monascus species and other host cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional promoters are used in Monascus species, then basic gene expression is maintained, but high expression of specific genes like acuF and Hsp cannot be achieved
Solution Approach 1:
The patent extracts and identifies specific promoter regions (SEQ ID NO: 1 and SEQ ID NO: 2) from the upstream regions of the acuF and Hsp genes. These promoter regions are then isolated and used in recombinant DNA constructs to achieve high expression of target proteins, separating the functional promoter elements from the rest of the genomic context.
Solution Approach 2:
The patent modifies promoter parameters by identifying specific nucleotide sequences with high expression potential. The promoter regions are characterized by specific sequence features that enable enhanced transcriptional activity, allowing transformation of normal gene expression into high-level protein production in host cells.
2Productivity
If promoter regions are identified and used in recombinant DNA technology, then protein over-expression is enabled, but the complexity of DNA manipulation increases
Solution Approach 1:
The patent segments the genomic DNA into functional components: the promoter region (SEQ ID NO: 1 or 2) and the coding region. This segmentation allows the promoter to be isolated, characterized, and reused in different recombinant constructs without requiring complete genomic sequences, simplifying the DNA manipulation process.
Solution Approach 2:
The identified promoter regions serve multiple functions: they drive expression of the native genes (acuF and Hsp) and can also be used in recombinant DNA technology to express various target proteins. This multi-functionality reduces the need for multiple different promoter elements, simplifying the overall system.
3Loss of information
If the Monascus EST database is searched for high expression genes, then useful gene information is obtained, but the time required for database analysis increases
Solution Approach 1:
The patent performs preliminary database searching and analysis to identify high-expression genes (acuF and Hsp) and their upstream promoter regions before actual protein production. This preliminary action retrieves valuable gene information in advance, allowing subsequent recombinant DNA construction to proceed more efficiently without repeated database queries.
Data Source
AI summary
Promoters and usage thereof. Two promoters, acuF and hsp promoters, comprise the nucleotide sequences of SEQ ID NO: 1 and 2, respectively.