Universal Detection Probes for Faster Multiplex PCR Quantification
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Solution Overview
Problem
Existing PCR detection methods using target-specific oligonucleotide probes are time-consuming and costly due to the need for individual design and validation for different target sequences, and they do not allow for efficient multiplex detection of multiple DNA targets.
Innovation Solution
A PCR detection system utilizing a universal detection probe with a 5'-universal tail that allows selective hybridization to a nucleotide sequence in the probe, eliminating the need for target-specific probe design and enabling faster assay development and cost savings.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If target-specific oligonucleotide probes are used for PCR detection, then detection precision and multiplex capability are improved, but time consumption and cost increase due to individual design and validation for each target sequence
Solution Approach 1:
The patent applies universality by designing a universal detection probe that can detect multiple different target sequences through combinatorial pairing with different primers. The universal probe contains a common detection region that hybridizes to a universal tail sequence appended to various target-specific primers, allowing a single probe design to serve multiple detection purposes across different targets, thereby eliminating the need for individual probe design and validation for each target sequence.
2Measurement precision
If target-specific oligonucleotide probes are used for PCR detection, then detection precision and multiplex capability are improved, but cost increases due to individual design and validation for each target sequence
Solution Approach 1:
The patent applies universality by designing a universal detection probe that can detect multiple different target sequences through combinatorial pairing with different primers. The universal probe contains a common detection region that hybridizes to a universal tail sequence appended to various target-specific primers, allowing a single probe design to serve multiple detection purposes across different targets, thereby eliminating the need for individual probe design and validation for each target sequence.
3Loss of time
If universal detection probe with 5'-universal tail is used, then time and cost are reduced by eliminating target-specific probe design, but probe complexity increases due to the universal tail sequence requirement
Solution Approach 1:
The patent applies segmentation by dividing the probe into two distinct functional segments: a universal detection region that remains constant and can be synthesized once, and a variable tail sequence region that is appended to different primers for different targets. This segmentation allows the complex universal detection probe to be constructed by combining a standardized detection module with simple tail sequences, reducing the overall complexity of probe development while maintaining detection capability across multiple targets.
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
The system reduces time and cost by allowing universal probe usage across different targets, facilitates multiplex detection, and enables the design of very short amplicons without unused stock, enhancing efficiency and versatility.
Implementation Method 1
the first 5'-universal tail of said first primer comprises a nucleotide sequence, the complement of which allows selective hybridization to a nucleotide sequence comprised in said first universal detection probe
Data Source
Figure 1~2
Figure 3A
Figure 3B
AI summary
The current invention relates to a PCR detection system comprising a reaction mixture for analyzing, quantitating or detecting a target nucleic acid, wherein the reaction mixture comprises: a. a first primer, wherein said first primer comprises a region configured to selectively hybridize to said target nucleic acid under amplification conditions and a first 5'-universal tail which is not complementary to the target nucleic acid sequence; b. a first universal detection probe, wherein said first universal detection probe comprises a first fluorophore, characterized in that the first 5'-universal tail of said first primer comprises a nucleotide sequence, the complement of which allows selective hybridization to a nucleotide sequence comprised in said first universal detection probe. The current invention further relates to methods and uses of aforementioned PCR detection system.