Lactobacillus casei Strain Screening for Fermented Milk Stability
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Solution Overview
Problem
Current methods for screening Lactobacillus casei fermentation agents for fermented milk beverages rely heavily on foreign imports and lack efficient screening techniques, leading to inconsistent acid production and reduced product stability due to excessive acidity and sucrose metabolism.
Innovation Solution
A method involving the screening of Lactobacillus casei strains with high-expression of the lac cluster gene and no enzymatic activity of β-fructosidase, using quantitative PCR to determine plasmid copy numbers and enzymatic activity, to selectively identify strains that rapidly metabolize lactose without sucrose metabolism, thereby controlling acidity and preventing over-sourness.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If Lactobacillus casei strains with high acid production rate are used for fermentation, then fermentation efficiency is improved, but excessive acidity occurs leading to over-sourness and reduced product stability
Solution Approach 1:
The patent changes the genetic parameters of Lactobacillus casei by introducing plasmids containing lac cluster genes (lacG, lacA, lacE, lacF, lacT) to enhance lactose metabolism capability. This genetic parameter modification enables controlled acid production from lactose, achieving high fermentation efficiency while preventing excessive acidity through regulated metabolic pathways.
Solution Approach 2:
The patent uses plasmids as intermediary carriers to transfer lac cluster genes into Lactobacillus casei strains. These plasmids act as mediators that enable controlled expression of lactose metabolism genes, allowing the bacteria to efficiently convert lactose to acid without producing excessive acidity that would harm product stability.
2Reliability
If foreign Lactobacillus casei fermentation agents are imported, then fermentation agent quality is improved, but dependency on imports increases and screening capability is reduced
Solution Approach 1:
The patent enables domestic self-service in fermentation agent development by establishing a complete screening system that identifies and characterizes local Lactobacillus casei strains. Through genetic analysis of lac cluster genes and metabolic pathway characterization, the method allows independent selection and optimization of fermentation agents without relying on foreign imports.
Solution Approach 2:
The patent performs preliminary characterization of Lactobacillus casei strains by analyzing plasmid copy numbers, lac cluster gene expression levels, and metabolic pathway activities before selecting fermentation agents. This preliminary screening enables identification of superior domestic strains that meet quality requirements, eliminating the need for post-import evaluation.
3Use of energy by moving object
If sucrose metabolism is active in Lactobacillus casei, then additional energy source is utilized, but post-acidification occurs leading to continued acidity increase and reduced shelf life
Solution Approach 1:
The patent extracts and analyzes specific metabolic pathway characteristics by measuring β-fructosidase activity and characterizing sucrose metabolism genes. Strains with active sucrose metabolism pathways are identified and excluded from selection, ensuring that chosen fermentation agents rely primarily on lactose metabolism, thereby preventing post-acidification and extending shelf life.
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 approach ensures consistent acid production, reduces post-acidification, and minimizes centrifugation sedimentation issues, resulting in improved product stability and shelf life of fermented milk beverages.
Implementation Method 1
the lactose enters cells through a gal-lac gene cluster of a phosphoenolpyruvate-lactose phosphotransferase system and is hydrolyzed into glucose and galactose
Implementation Method 2
Sucrose is transferred into the cells through a phosphoenolpyruvate-sucrose phosphotransferase system
Implementation Method 3
The phosphorylated sucrose is hydrolyzed by β-fructosidase (EC 3.2.1.26) in the cells to generate glucose-6-phosphate and fructose
Implementation Method 4
the generated galactose is decomposed into organic acids under the action of a gene cluster galR-galKTEM
Implementation Method 5
amplifying tuf and lac cluster genes, using the tuf gene as a control, and using an equation Nrelatives=(1+E)−ΔCT to determine a plasmid copy number of the lac cluster gene
Data Source
AI summary
A method for screening Lactobacillus casei fermentation agent is provided. The method is specifically implemented by screening a Lactobacillus casei strain with high-expression of a lac cluster gene and no enzymatic activity of β-fructosidase. The Lactobacillus casei strain that rapidly metabolizes lactose is screened through high expression of the lac cluster gene, such that the problem of large differences in acid production in fermented milk beverage products is avoided.