ATPS Purification of Short Nucleic Acid Fragments
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Solution Overview
Problem
Current methods are inadequate for efficiently isolating, purifying, and concentrating short nucleic acid fragments less than 250 base pairs, which are crucial for applications like liquid biopsy, due to low yield and diagnostic sensitivity issues.
Innovation Solution
The use of an aqueous two-phase system (ATPS) comprising specific polymers and salts allows for the isolation, concentration, and purification of short nucleic acid fragments by partitioning them into distinct phases, thereby enhancing yield and purity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If conventional nucleic acid extraction methods are used, then general nucleic acids can be isolated, but short nucleic acid fragments less than 250 bp cannot be efficiently isolated with sufficient yield and purity
Solution Approach 1:
The invention changes the physical-chemical parameters of the extraction system by using aqueous two-phase systems with specific polymer-salt combinations (PEG 8000-PEG 1000, PEG 6000-PES 300, PEG 4000-PEG 1000) at controlled temperatures and pH levels, enabling selective partitioning of short nucleic acid fragments less than 250 bp into one phase while excluding longer fragments and contaminants, thereby achieving both high purity and sufficient yield
Solution Approach 2:
The invention exploits phase transition phenomena in aqueous two-phase systems where the addition of polymers and salts to biological samples causes the system to separate into two immiscible aqueous phases, with short nucleic acid fragments preferentially partitioning into one phase, enabling their isolation from longer fragments and contaminants through simple phase separation
2Measurement precision
If multiple purification steps are performed to achieve high purity of short nucleic acid fragments, then diagnostic sensitivity improves, but processing time and complexity increase
Solution Approach 1:
The invention merges multiple purification functions (isolation, concentration, and purification of short nucleic acid fragments) into a single aqueous two-phase system extraction step, eliminating the need for sequential purification steps while achieving the purity required for diagnostic sensitivity in downstream applications like PCR and sequencing
3Ease of manufacture
If conventional extraction kits are used, then general nucleic acid extraction can be performed, but short nucleic acid fragments less than 250 bp are lost or contaminated
Solution Approach 1:
The invention modifies the extraction parameters by using aqueous two-phase systems with specific polymer-salt combinations (PEG 8000-PEG 1000, PEG 6000-PES 300, PEG 4000-PEG 1000) at controlled temperatures and pH levels, enabling selective partitioning of short nucleic acid fragments less than 250 bp into one phase while excluding longer fragments and contaminants, thereby achieving both high purity and sufficient yield
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 achieves high purity and concentration of short nucleic acid fragments, improving diagnostic sensitivity and specificity, and is compatible with downstream applications like PCR and sequencing without additional purification steps.
Implementation Method 1
The use of an aqueous two-phase system (ATPS) comprising specific polymers and salts allows for the isolation, concentration, and purification of short nucleic acid fragments by partitioning them into distinct phases
Implementation Method 2
The use of an aqueous two-phase system (ATPS) comprising specific polymers and salts allows for the isolation, concentration, and purification of short nucleic acid fragments by partitioning them into distinct phases, thereby enhancing yield and purity
Data Source
AI summary
The present invention relates to methods of using aqueous two-phase system (ATPS) for the isolation, concentration and/or purification of short nucleic acid fragment having about or less than 250 base pairs (bp). In one embodiment, the present invention provides a composition and kit for the purification of short nucleic acid fragments having about or less than 250 base pairs from nucleic acid-containing biological materials. In another embodiment, the present invention provides uses of certain salts and/or polymers in a two-phase system for the purification of short nucleic acid fragments having about or less than 250 base pairs from nucleic acid containing biological materials.