Target reporter constructs and uses thereof
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Solution Overview
Problem
Current nucleic acid detection technologies, particularly for cell-free miRNAs, are limited by their high cost, complexity, and inability to efficiently quantify and interpret minute populations, making them impractical for widespread diagnostic use.
Innovation Solution
Target reporter constructs (TRCs) are developed, comprising a target sequence, bridge sequence, and accessory sequence, which undergo rolling circle replication (RCR) and amplification (RCA) to detect and quantify nucleic acids, allowing for multiplex detection and analysis on a single platform.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If current nucleic acid detection technologies (RT-PCR, NGS, microarray) are used, then detection sensitivity and accuracy are improved, but cost and complexity increase significantly
Solution Approach 1:
The detection system is segmented into distinct functional modules: target reporter constructs (TRCs) for specific nucleic acid detection, rolling circle replication (RCR) for amplification, and rolling circle amplification (RCA) for signal generation. Each module performs a specific function, allowing the system to achieve high sensitivity through specialized mechanisms while reducing overall system complexity through functional decomposition.
Solution Approach 2:
The patent introduces target reporter constructs (TRCs) as intermediary molecules that bridge the target nucleic acid and the detection system. These TRCs contain complementary sequences that hybridize to target nucleic acids, initiating RCR and RCA reactions. This intermediary approach simplifies the detection process by creating a dedicated recognition and amplification pathway that is easier to implement than direct detection methods.
2Measurement precision
If current nucleic acid detection technologies are used, then detection accuracy is improved, but cost increases significantly
Solution Approach 1:
The patent employs disposable target reporter constructs (TRCs) and consumable reagents for RCR and RCA reactions. These single-use components are designed to be inexpensive to manufacture, allowing high-accuracy detection without the need for expensive reusable equipment or complex infrastructure. The disposable nature of these components eliminates costly maintenance, calibration, and sterilization requirements associated with traditional detection systems.
3Measurement precision
If traditional detection methods are used for minute nucleic acid populations, then detection sensitivity is improved, but the ability to efficiently quantify and interpret results deteriorates
Solution Approach 1:
The patent transforms the detection output from qualitative to quantitative by measuring physical parameters such as fluorescence intensity, absorbance, or particle count generated during RCR and RCA reactions. These parameter changes provide a direct, proportional relationship between the amount of target nucleic acid and the measured signal, enabling efficient quantification and interpretation of minute nucleic acid populations without complex analysis procedures.
4Device complexity
If single-pathogen analysis systems are used, then simplicity is maintained, but the need for additional tests for each target increases
Solution Approach 1:
The patent designs a universal detection platform that can identify multiple different nucleic acid targets using the same basic system architecture. Different target reporter constructs (TRCs) with complementary sequences specific to various pathogens can be used with the same RCR and RCA machinery. This multi-functional approach allows simultaneous or sequential detection of multiple pathogens without requiring separate testing systems, thereby reducing the time and resources needed for comprehensive diagnostic analysis.
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
TRCs provide rapid, cost-effective, and sensitive detection of nucleic acids, including miRNAs, with high specificity and scalability, reducing technical requirements and enabling widespread diagnostic applications.
Implementation Method 1
The TRCs undergo rolling circle replication (RCR) and amplification (RCA) to detect and quantify nucleic acids
Implementation Method 2
The TRCs undergo rolling circle replication (RCR) and amplification (RCA) to detect and quantify nucleic acids
Implementation Method 3
a target sequence complementary to the target nucleic acid
Data Source
AI summary
Provided herein are methods and compositions for the detection of target nucleic acids using target reporter constructs (TRCs) which comprise target sequences complementary to the target nucleic acid. Further provided are methods of replicating the TRCs using rolling circle replication and/or rolling circle amplification to produce replicated TRCs which can be detected using probe sequences within the replicated TRCs.