Transmembrane DNA Sensor for Lysis-Free Intracellular RNA Detection
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Solution Overview
Problem
Current methods for non-invasive detection of nucleic acid targets in live cells are limited by the lack of suitable tools, leading to disruptive sample processing and the exclusion of rare cell populations with unique transcriptomic profiles.
Innovation Solution
A DNA sensor with a hydrophobic tag that facilitates insertion through the lipid bilayer membrane using a flip-flop mechanism, enabling lysis-free detection of nucleic acids within lipid vesicles by leveraging toehold-mediated strand displacement and DNA hybridization design.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional detection methods are used, then nucleic acid targets can be detected, but cell lysis is required which disrupts the sample and excludes rare cell populations
Solution Approach 1:
The sensor is divided into distinct functional segments: a hydrophobic tag for membrane insertion, a toehold domain for target binding, and a reporter domain for signal generation. This segmentation allows the sensor to perform detection functions while the hydrophobic tag handles membrane interaction, enabling lysis-free detection
Solution Approach 2:
The hydrophobic tag acts as an intermediary that facilitates sensor insertion through the lipid bilayer membrane. This intermediary component enables the sensor to cross the membrane barrier without disrupting the cell, allowing intracellular target detection while maintaining cell integrity
2Measurement precision
If cell lysis is performed for nucleic acid quantification, then detection can be achieved, but rare cell populations with unique transcriptomic profiles are lost
Solution Approach 1:
The sensor performs detection functions autonomously within the cell without requiring external processing or lysis. The sensor self-assembles, inserts into the membrane, binds to intracellular targets, and generates detectable signals, enabling reliable analysis of rare cell populations while maintaining cell viability
Solution Approach 2:
The sensor utilizes changes in conformational state upon target binding to generate detectable signals. The transition from unbound to bound states produces measurable optical or electrical signals, enabling quantification without cell disruption and preserving rare cell populations for further analysis
3Ease of operation
If a transmembrane sensor is designed with hydrophobic tag, then membrane insertion is facilitated, but sensor design complexity increases
Solution Approach 1:
The hydrophobic tag is pre-attached to the sensor molecule during synthesis, enabling spontaneous membrane insertion when the sensor encounters the lipid bilayer. This preliminary preparation eliminates the need for complex insertion protocols or additional components, simplifying the overall operation despite the enhanced design
Solution Approach 2:
The sensor combines hydrophobic tag molecules with nucleic acid sensing domains to create a composite structure. This composite design integrates membrane-interacting properties with specific target recognition capabilities, facilitating membrane insertion while maintaining detection function through material composition rather than complex structural arrangements
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 sensor allows for non-invasive detection of intracellular nucleic acids, reducing sample disruption and enabling faster, more efficient analysis of nucleic acids in biological samples.
Implementation Method 1
A DNA sensor with a hydrophobic tag that facilitates insertion through the lipid bilayer membrane using a flip-flop mechanism
Implementation Method 2
enabling lysis-free detection of nucleic acids within lipid vesicles by leveraging toehold-mediated strand displacement and DNA hybridization design
Data Source
AI summary
The present disclosure provides compositions and methods related to nucleic acid sensors. In particular, the present disclosure provides nucleic acid sensors that can span through lipid bilayer membranes to detect internal nucleic acid targets present in vesicles.