DNA Strand Luminophore Positioning for Binding Site Localization
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Solution Overview
Problem
Current methods for determining the precise binding location of reagents to DNA strands are limited by calibration accuracy and are prone to off-target binding due to physical and environmental factors, making it challenging to focus imaging systems effectively for optimal localization.
Innovation Solution
A method involving a DNA strand with luminophores covalently attached at predetermined positions, allowing for precise determination of focus positions and reaction sites using optical detectors and manipulators, thereby improving localization accuracy and reducing risks of luminophore detachment or relocation.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If optical tweezers combined with fluorescence microscopy are used to study biomolecular processes, then valuable information about reagent-DNA interactions can be obtained, but the accuracy of determining binding locations is limited by calibration accuracy and system factors
Solution Approach 1:
The patent introduces luminophores as intermediary markers covalently attached to the DNA strand at predetermined positions. These luminophores serve as reliable reference points that are independent of system calibration, allowing accurate determination of binding locations through optical detection without depending on calibration accuracy of trap position, bead sizes, or DNA tension.
Solution Approach 2:
The patent replaces the mechanical calibration system (relying on trap position, bead sizes, and DNA tension calibration) with an optical reference system using covalently attached luminophores. This substitution eliminates the need for mechanical calibration and provides more reliable and accurate binding location determination.
2Measurement precision
If in-situ labelling methods are used to mark specific sites on DNA molecules, then binding sites can be identified, but extra incubation steps and specific buffer conditions are required which may cause off-target binding
Solution Approach 1:
The patent applies preliminary action by covalently attaching luminophores to the DNA strand at predetermined positions before the actual binding experiment. This pre-labeling approach eliminates the need for post-experiment incubation steps and avoids the complexity of in-situ labelling procedures, while ensuring consistent and reliable reference points for binding location determination.
3Quantity of substance
If multiple binding sites for enzymes are used as markers, then coverage can be improved, but the stochastic distribution of labelling sites creates uncertainty in position determination
Solution Approach 1:
The patent applies local quality by placing luminophores at specific predetermined positions along the DNA strand rather than using stochastic enzyme binding sites. This ensures that each luminophore serves as a known reference point at a determined location, eliminating the uncertainty associated with stochastic labelling while maintaining sufficient coverage for accurate position determination.
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 approach enhances the reliability of determining reagent binding sites on DNA strands with high precision, independent of system calibrations, and reduces off-target binding, enabling more accurate biomolecular process studies.
Implementation Method 1
providing a DNA strand comprising one or more luminophores attached to the strand at one or more predetermined positions along the strand
Implementation Method 2
trapping and/or manipulating the strand by manipulating the handles in a trapping setup
Data Source
AI summary
A method of detecting a biomolecular process is provided. The method comprises: providing a DNA strand (11) comprising one or more luminophores (3) attached to the strand (11) at one or more predetermined positions along the strand; attaching handles (7A, 7B) to the DNA strand; and trapping and/or manipulating the strand (11) by manipulating the handles (7A, 7B) in a trapping setup. The method also comprises determining a position of at least one of the one or more luminophores (3) with respect to at least part of the trapping setup and determining on the basis of the one or more luminophores (3) one or more of: a focus position of at least part of an optical detector and/or an illumination system; a focus position of at least part of an optical manipulator; a position of at least part of the strand (11); a position of a portion of the strand (11) relative to one or more other portions of the strand (11); a reaction or binding position on the strand (11) of a reagent interacting with the strand (11). The one or more luminophores (3) are covalently attached to the DNA strand. An associated DNA strand is also provided.