Ultrafiltration of Duplex Nucleotides for Fast Purity Separation
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Solution Overview
Problem
Manufacturing of oligonucleotides is time and resource intensive, and there is a need for improved methods and apparatuses to enhance the purity and separation of double-stranded and single-stranded nucleic acids.
Innovation Solution
The use of ultra-filtration methods and apparatuses that selectively retain double-stranded nucleic acids while allowing single-stranded nucleic acids to permeate, utilizing filtration elements with specific pore sizes and molecular weight cutoffs to achieve separation and purification.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If traditional oligonucleotide manufacturing methods are used, then production can be maintained, but manufacturing time and resource consumption are excessive and purity is insufficient
Solution Approach 1:
The patent employs ultrafiltration membranes with specific pore size cutoffs (e.g., 100 kDa, 50 kDa, 10 kDa) to separate double-stranded nucleic acids from single-stranded contaminants. The porous membrane structure allows selective passage of molecules based on size, enabling efficient purification without time-consuming chromatography steps.
Solution Approach 2:
The invention changes the physical parameters of the separation process by using ultrafiltration with defined molecular weight cutoffs. By selecting appropriate membrane cutoff values (e.g., 100 kDa for retaining dsDNA while allowing ssDNA to pass), the method achieves rapid purification in a single step, dramatically reducing manufacturing time compared to traditional multi-step protocols.
2Manufacturing precision
If traditional purification methods are used, then separation can be achieved, but the process is complex and resource intensive
Solution Approach 1:
The patent extracts and utilizes the size-based separation property of nucleic acids by passing the mixture through ultrafiltration membranes. This single extraction step using differential permeability replaces complex multi-step purification schemes, simplifying the overall process while maintaining high separation efficiency.
Solution Approach 2:
Ultrafiltration membranes with controlled pore sizes serve as the core separation medium. These porous materials provide size-selective filtration in a single device, eliminating the need for multiple purification components and reducing apparatus complexity while achieving effective separation of dsDNA from ssDNA contaminants.
3Productivity
If conventional manufacturing approaches are used, then production can continue, but resource consumption and cost are excessive
Solution Approach 1:
The ultrafiltration process enables continuous purification where the filtrate can be collected and further processed or used directly. The membrane filtration operates continuously without interruption, maintaining productive flow and eliminating idle time between purification steps, thereby improving manufacturing efficiency and reducing resource waste.
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
Enhances the purity of double-stranded nucleic acids and separates them effectively from single-stranded nucleic acids, improving manufacturing efficiency and product quality.
Implementation Method 1
subjecting the mixture to an ultra-filtration step that selectively retains the double-stranded nucleic acid comprising the hybridized first and second single-stranded nucleic acids but not the single-stranded nucleic acid(s)
Implementation Method 2
a membrane with a molecular weight (MW) cutoff that is at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, or 80% less than the MW of the double-stranded nucleic acid
Data Source
AI summary
Disclosed herein are methods of purifying oligonucleotides, for example, by separating double-stranded oligonucleotides from single-stranded oligonucleotides. Also provided are apparatuses for performed such methods. Also provided are double-stranded nucleic acid compositions and single-stranded nucleic acid compositions made using the methods and apparatuses disclosed herein. Also provided are methods of producing circular double-stranded nucleic acids.