SgTPS7 Germacrene Synthase Expression for High-Purity Germacrene
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Solution Overview
Problem
Current methods for extracting germacrene from plant tissues are limited by low yield and purity, necessitating a more efficient production method.
Innovation Solution
Isolation of the germacrene synthase SgTPS7 gene and protein from Sindora glabra Merr. ex de Wit, which catalyzes the production of germacrene using Escherichia coli, through the use of a pET30a-SgTPS7 recombinant plasmid for expression and purification.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of manufacture
If germacrene is extracted from plant tissues, then the extraction process is simple, but the yield and purity are greatly limited
Solution Approach 1:
The patent replaces the mechanical extraction process with a biological synthesis system. The germacrene synthase gene (SgTPS7) is cloned and expressed in Escherichia coli, which then catalyzes the conversion of farnesyl pyrophosphate (FPP) to germacrene D. This substitution of mechanical extraction with biological catalysis resolves the contradiction by achieving both high productivity (through bacterial mass cultivation and high enzyme expression) and high purity (through specific enzymatic catalysis), while maintaining ease of manufacture through established recombinant protein expression protocols.
Solution Approach 2:
The patent changes the fundamental parameter of germacrene production from extraction-based to synthesis-based. By cloning the SgTPS7 gene and expressing it in E. coli, the system transforms the production approach from obtaining germacrene through physical extraction from plant tissues to generating it through enzymatic catalysis in a controlled biological system. This parameter change enables high-yield production with high purity while maintaining process simplicity through standard molecular biology techniques.
2Productivity
If genetic engineering methods are used to produce germacrene using Escherichia coli, then the productivity and purity are improved, but the device complexity increases
Solution Approach 1:
The patent segments the complex genetic engineering process into distinct, manageable modules: (1) gene cloning of SgTPS7 from Sindora glabra, (2) construction of recombinant expression vector, (3) transformation into E. coli competent cells, (4) protein expression induction, (5) cell lysis and enzyme extraction, and (6) catalytic reaction. This segmentation allows each step to be optimized and controlled independently, reducing overall process complexity while maintaining high productivity and purity through precise control at each stage.
Solution Approach 2:
The patent introduces several intermediary elements that simplify the genetic engineering process: a plasmid vector as an intermediary carrier for the SgTPS7 gene, IPTG as an intermediary inducer for protein expression, and FPP as an intermediary substrate for the enzymatic reaction. These intermediaries serve as standardized, well-characterized components that bridge different stages of the process, making the overall system more controllable and less complex despite the multi-step nature of genetic engineering.
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
Enables the mass production of germacrene with improved yield and purity by utilizing genetic engineering techniques.
Implementation Method 1
The gene encodes the protein germacrene synthase D, which can catalyze the production of germacrene
Data Source
AI summary
The present disclosure discloses a germacrene synthase SgTPS7 protein, a coding gene and a use in synthesizing germacrene, relating to the technical field of germacrene synthesis of Sindora glabra Merr. ex de Wit. The present disclosure isolates a gene of germacrene synthase D from the genome of Sindora glabra Mer. ex de Wit. The gene encodes the protein germacrene synthase D which can catalyze the production of germacrene and can be used for the production and application of germacrene.