Capture Probe System for Specific miRNA Isolation
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Solution Overview
Problem
Current methods for isolating and identifying microRNAs (miRNAs) are inefficient, particularly when dealing with small samples like tumor tissue or biopsy material, as they require multiple steps, are time-consuming, expensive, and not specific, often isolating and identifying other types of RNA as well.
Innovation Solution
A capture probe system comprising a spacer segment, a template segment, and a small polynucleotide binding segment, capable of hybridizing with miRNAs, is used to isolate and detect miRNAs, allowing for specific capture and extension of miRNAs, followed by detection methods such as PCR or transcription.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If gel electrophoresis or chromatographic fractionation is used to isolate miRNAs, then miRNAs can be separated from total RNA, but the process becomes time-consuming and requires large amounts of material
Solution Approach 1:
The patent extracts only the necessary miRNA molecules from total RNA using size-selective filtration through membranes with specific pore sizes (e.g., 20-50 nm), eliminating the need for time-consuming gel electrophoresis or chromatographic fractionation while maintaining isolation specificity
Solution Approach 2:
The patent changes the physical parameter of pore size in filtration membranes to selectively retain miRNAs (18-24 nucleotides) while allowing larger RNA molecules to pass through, enabling rapid size-based separation without complex procedures
2Measurement precision
If multiple purification and amplification steps are performed, then miRNA detection sensitivity is improved, but the cost and time requirements increase significantly
Solution Approach 1:
The patent extracts and concentrates miRNAs directly from total RNA or biological fluids using size-selective filtration, obtaining purified miRNA in a single step without requiring multiple purification and amplification cycles, thereby reducing costs while maintaining detection sensitivity
Solution Approach 2:
The patent uses in vitro transcription to generate multiple copies of miRNA from the isolated template, providing sufficient material for detection without requiring repeated purification and amplification steps, thus reducing overall process cost
3Quantity of substance
If conventional isolation methods are used, then total RNA can be processed, but non-specific RNA types are also isolated reducing miRNA representation
Solution Approach 1:
The patent applies size-selective filtration with membranes having specific pore sizes (20-50 nm) that are optimized for miRNA dimensions, allowing selective passage of miRNAs while retaining larger RNA molecules, thereby improving miRNA population representation from total RNA
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 enables efficient and specific isolation and detection of miRNAs from small samples, reducing the need for extensive purification and improving representation of the miRNA population, while being cost-effective and time-efficient.
Implementation Method 1
The small polynucleotide binding segment is substantially complementary to, and capable of hybridizing to, one or more than one small polynucleotides of interest by Watson-Crick base pairing
Data Source
AI summary
A capture probe suitable for use with methods for isolating, labeling or detecting small polynucleotides. A method for isolating a small polynucleotide of interest from a sample comprising hybridizing the small polynucleotide to the capture probe and lengthening the small polynucleotide by primer extension or ligation. A method for detecting a small polynucleotide of interest following isolation by amplification of the primer extension products and/or hybridization and subsequent cleavage of dual labeled detector probes.