Bis-Boron Fluorescent Dyes for Stable Multiplex Sequencing
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Solution Overview
Problem
Existing fluorescent dyes for nucleic acid sequencing face challenges such as poor chemical stability in aqueous environments, difficulty in achieving distinguishable spectral properties, and compatibility with reagent chemistries, limiting their use in high-throughput sequencing methods.
Innovation Solution
Development of fused tetracyclic bis-boron containing heterocycles with improved chemical stability and tunable absorption/emission properties, suitable for blue light excitation, allowing for strong fluorescence and compatibility with sequencing reagents.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional fluorescent dyes are used for nucleic acid sequencing, then detection can be performed, but the dyes exhibit poor chemical stability in aqueous environments and limited shelf life
Solution Approach 1:
The patent modifies the chemical structure of fluorescent dyes by incorporating boron-containing heterocyclic rings and specific substituents (such as fluoro, chloro, alkoxy groups) to enhance photo-stability and chemical stability in aqueous environments. This structural parameter change directly addresses the reliability and shelf life issues of conventional dyes.
Solution Approach 2:
The invention creates composite fluorescent dye molecules that combine boron-containing heterocyclic structures with various aromatic systems and substituents. These composite structures provide both the desired fluorescence properties and improved stability, resolving the contradiction between detectability and chemical stability.
2Productivity
If multiple fluorescent dyes are used for multiplex detection, then throughput increases, but it becomes difficult to find dyes with substantially resolved absorption and emission spectra
Solution Approach 1:
The patent designs fluorescent dyes with specific local structural features (different substituents, heterocyclic variations) that tune the absorption and emission spectra of individual dyes. This allows creation of multiple dyes with distinct spectral characteristics suitable for multiplex detection while maintaining comparable excitation efficiencies.
Solution Approach 2:
By systematically varying structural parameters of the boron-containing heterocyclic dyes (such as substituent types, ring positions, conjugation patterns), the patent generates a family of dyes with resolved spectra that can be excited by the same or similar wavelengths, enabling multiplex detection without sacrificing spectral resolution.
3Illumination intensity
If high power light sources are used for excitation, then fluorescence signal strength increases, but the dyes must have sufficient photo-stability to withstand such excitation
Solution Approach 1:
The patent enhances photo-stability by incorporating boron-containing heterocyclic structures with specific substituents (fluoro, chloro, alkoxy groups) that protect the dye molecules from photodegradation. This allows the dyes to withstand high power excitation sources while maintaining stable fluorescence signals.
Solution Approach 2:
The invention develops dyes that are stable enough for repeated use with high power lasers, eliminating the need for frequent replacement and ensuring consistent performance throughout the sequencing process.
4Measurement precision
If fluorescent dyes are optimized for strong fluorescence signals, then sequencing accuracy improves, but compatibility with reagent chemistries such as DNA synthesis solvents, buffers, and enzymes may be compromised
Solution Approach 1:
The patent modifies dye structures to include substituents and linkers that enhance solubility in aqueous buffers and compatibility with DNA synthesis reagents while maintaining strong fluorescence. The boron-containing heterocyclic core provides a platform for adding such functional groups without compromising optical properties.
Solution Approach 2:
The invention separates the optical function (provided by the boron-heterocyclic core) from the chemical compatibility function (provided by peripheral substituents and linkers). This allows optimization of each aspect independently, achieving both strong fluorescence and reagent compatibility.
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 new dyes provide enhanced fluorescence intensity and stability, enabling better optical resolution and reduced sequencing errors, facilitating high-throughput nucleic acid sequencing.
Implementation Method 1
Fluorescent dyes containing fused tetracyclic bis-boron heterocycle and their uses in sequencing
Implementation Method 2
tunable absorption/emission properties, suitable for blue light excitation
Data Source
AI summary
The present application relates to dyes containing fused tetracyclic bis-boron containing heterocycle and their uses as fluorescent labels. These dyes may be used as fluorescent labels for nucleotides in nucleic acid sequencing applications.