Nucleic Acid Isolation from Water Using Alginate Gel Concentration
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Solution Overview
Problem
Current methods for isolating nucleic acids from water samples are time-consuming, labor-intensive, and inefficient, often losing biomolecules due to separation of sediment and liquid phases, and require expensive equipment like ultracentrifuges.
Innovation Solution
A method involving centrifugation to separate sediment and liquid phases, followed by the use of alginate derivatives to form gels in the liquid phase, concentrating biomolecules, and subsequent lysis and extraction using commercially available reagents, allowing simultaneous isolation from both fractions without the need for ultracentrifugation.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If centrifugation is used to remove solids from water samples, then the liquid phase is purified for analysis, but biomolecules bound to solids are lost in the sediment
Solution Approach 1:
The patent combines the sediment fraction and liquid phase fraction into a single processing stream. After centrifugation separates the sample into sediment and supernatant, both fractions are processed together through the nucleic acid extraction protocol, ensuring that biomolecules from both sediment-bound and free-floating sources are recovered simultaneously, eliminating the loss associated with discarding either fraction
Solution Approach 2:
The patent segments the water sample into two distinct fractions (sediment and liquid phase) through centrifugation, then processes each fraction separately through lysis and nucleic acid extraction before combining the results. This segmentation allows targeted optimization for each fraction type while ensuring comprehensive biomolecule recovery
2Quantity of substance
If conventional nucleic acid extraction methods are used on large volume water samples, then sufficient sample volume is available for analysis, but the extraction becomes time-consuming and labor-intensive
Solution Approach 1:
The patent extracts and concentrates nucleic acids from large volume water samples (up to 200 ml) into a small final volume suitable for downstream molecular analysis. The method efficiently removes water and other contaminants while concentrating the target nucleic acids, transforming a large-volume problem into a small-volume solution that is immediately suitable for PCR and other molecular techniques
Solution Approach 2:
The patent employs parameter changes in the form of pH adjustment and chaotropic salt addition to facilitate rapid nucleic acid binding to silica membranes. By changing the chemical parameters of the solution (adding guanidinium thiocyanate and adjusting pH), the method enables fast, efficient nucleic acid capture and concentration from large volumes without requiring lengthy processing steps
3Quantity of substance
If ultracentrifugation or ultrafiltration is used to concentrate biomolecules from large-volume samples, then virus enrichment is achieved, but expensive specialized equipment is required
Solution Approach 1:
The patent replaces expensive, complex ultracentrifugation and ultrafiltration equipment with simple, disposable silica-based spin columns. These inexpensive columns perform the concentration and purification function that would otherwise require sophisticated instrumentation, making the method accessible to routine laboratories without specialized equipment
Solution Approach 2:
The patent substitutes mechanical concentration methods (ultracentrifugation, ultrafiltration) with a chemical-biological approach using silica membrane binding. Instead of relying on mechanical forces to concentrate biomolecules, the method uses chemical interactions between nucleic acids and silica surfaces in the presence of chaotropic salts, eliminating the need for expensive mechanical concentration devices
4Quantity of substance
If NaCl/PEG precipitation is used to concentrate viruses, then virus particles are enriched, but the precipitates are difficult to dissolve and further processing is complex
Solution Approach 1:
The patent extracts nucleic acids directly from the water sample matrix using silica-based columns, bypassing the formation of difficult-to-process precipitates entirely. By taking out the nucleic acids in a soluble, column-bound form rather than as precipitated virus particles, the method eliminates the dissolution and re-suspension steps that complicate PEG/NaCl methods
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 method significantly increases the sensitivity of nucleic acid detection by ensuring no loss of biomolecules and can be completed within an hour, using standard equipment, enabling efficient extraction from both sedimentable and nonsedimentable biomolecules in water samples.
Implementation Method 1
separating the sample into a sediment fraction and a liquid phase
Implementation Method 2
the use of polysaccharide derivatives for complexing the biomolecules contained in a sample
Data Source
AI summary
The invention relates to a simple and very rapid method for simultaneously isolating nucleic acids from sedimentable and nonsedimentable biomolecules in water samples. The water samples can be, inter alia, drinking water, industrial water, surface water or else wastewater. The biomolecules are bacteria, bacteriophages, protozoa, viruses, and free-circulating nucleic acids (e.g., plasmid DNA) encoding antibiotic resistances. According to the invention, a polysaccharide derivative is added to the water sample and the biomolecules am thus enriched. After dissolution of the pellet and lysis of the biomolecules, the nucleic acids are bound to a solid phase, purified and eluted in a known manner.