Phi6 Bacteriophage RNA Controls for Thermostable Nucleic Acid Detection
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Solution Overview
Problem
Existing nucleic acid amplification methods require heat-resistant and ribonuclease-resistant RNA standards that can withstand high temperatures without degrading, as current armored RNA constructs degrade rapidly at elevated temperatures, necessitating cold storage which is costly.
Innovation Solution
A composition comprising Phi6 bacteriophage suspended in a buffer with a pH between 7-9, including a chelating agent and preservative, optionally with additional components like BSA, gelatin, poly rA RNA, glycerol, amino acids, or polyethylene glycol, providing thermostability up to 110 days at 30°C and 28 days at 45°C.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If armored RNA constructs are used as internal control nucleic acids, then they can reflect the properties of target RNA nucleic acids, but they degrade rapidly at elevated temperatures requiring costly cold storage
Solution Approach 1:
The patent changes the physical and chemical parameters of the RNA standard by encapsulating it within Phi6 bacteriophage particles. This encapsulation fundamentally alters the stability parameters of the RNA, enabling it to withstand temperatures up to 45°C for 28 days and 30°C for 110 days, thereby eliminating the need for costly cold storage while maintaining reliability as an internal control
Solution Approach 2:
The patent creates a composite structure where the RNA standard is encapsulated within Phi6 bacteriophage particles. This composite construction combines the RNA core with the protective viral capsid and envelope, forming a thermostable armored particle that resists degradation at elevated temperatures while retaining the RNA's functional properties for use as an internal control
2Measurement precision
If RNA is used as internal control nucleic acid to reflect target nucleic acid properties, then detection accuracy improves, but RNA degradation due to inherent sensitivity to alkaline pH and ribonucleases occurs
Solution Approach 1:
The patent applies beforehand cushioning by pre-encapsulating the RNA standard within Phi6 bacteriophage particles before use. The viral capsid and envelope act as protective barriers that cushion the RNA against harmful factors such as ribonucleases and alkaline pH conditions, preventing degradation while maintaining the RNA's detection accuracy functionality
Solution Approach 2:
The Phi6 bacteriophage structure serves as an intermediary between the RNA standard and the harsh environmental conditions. The viral capsid and envelope act as mediating layers that protect the RNA from direct exposure to ribonucleases and alkaline pH, allowing the RNA to maintain its integrity and measurement precision without direct contact with harmful factors
3Stability of the object's composition
If cold storage is used to maintain RNA standard stability, then degradation is prevented, but storage costs increase
Solution Approach 1:
The patent changes the stability parameters of the RNA standard through encapsulation in Phi6 bacteriophage, shifting the temperature stability parameter from requiring cold storage (4°C or lower) to tolerating elevated temperatures (30°C for 110 days, 45°C for 28 days). This parameter change eliminates the need for energy-intensive cold storage while maintaining composition stability, thereby reducing storage costs
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 Phi6 bacteriophage composition maintains stability and functionality for extended periods at elevated temperatures, allowing for reliable nucleic acid detection methods without the need for cold storage, thereby reducing costs and ensuring assay integrity.
Implementation Method 1
heat-resistant, ribonuclease-resistant RNA standard formulations
Implementation Method 2
including a chelating agent and a preservative
Data Source
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AI summary
The disclosure provides control compositions used in nucleic acid amplification. Various modifications to the disclosed compositions and methods of using the same, devices, and kits are described.