Chromosomal Inversion Detection via Strand-Specific Probes
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Solution Overview
Problem
Current methods for detecting chromosomal inversions are insensitive, leading to underestimation of their significance in diseases, including cancer, as they cannot accurately identify inversions due to limitations in banding patterns and whole-chromosome painting techniques.
Innovation Solution
The development of a method using single-stranded probes that target specific chromatids, allowing for the detection of inversions by hybridizing labeled probes to sister chromatids, which change their hybridization pattern in the presence of an inversion, enabling visualization of inverted DNA sequences.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If whole-chromosome painting by FISH is used, then chromosome identification is achieved, but inversion detection sensitivity is insufficient
Solution Approach 1:
The invention segments the chromosome into individual chromatids by using strand-specific probes that target only one strand of the DNA double helix. This allows differentiation between the two sister chromatids, enabling detection of inversions that affect only one chromatid. The segmentation is achieved through CO-FISH methodology that separates and labels specific strands, providing resolution at the chromatid level rather than whole-chromosome level.
Solution Approach 2:
The invention applies local quality by using strand-specific probes that are complementary to specific sequences on one chromatid but not the other. This creates localized labeling patterns that highlight specific regions on particular chromatids, allowing detection of localized inversions and structural abnormalities without requiring analysis of the entire chromosome.
2Measurement precision
If G- or R-banding is used, then banding pattern changes can be detected, but small inversions with breakpoints near midpoints are difficult to detect
Solution Approach 1:
The invention replaces the mechanical banding pattern visualization method with a molecular probe-based detection system. Instead of relying on physical banding patterns that may not resolve small inversions, the method uses fluorescently labeled strand-specific probes that bind to specific DNA sequences. This substitution enables detection at the nucleotide level, making small inversions and breakpoints visible regardless of their location on the chromosome.
3Productivity
If standard karyotype analysis is used, then gross chromosomal aberrations can be identified, but inversions are largely missed
Solution Approach 1:
The invention adds a new dimension to chromosomal analysis by introducing strand-specificity as a detectable feature. Traditional karyotype analysis looks at chromosome structure in two dimensions (banding patterns, chromosome size), but this method adds a third dimension - the directional orientation of DNA strands. By labeling only one strand with fluorescent probes, the method creates asymmetric labeling patterns that reveal inversions and other structural abnormalities that are invisible to conventional methods.
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 provides a sensitive and accurate method for detecting chromosomal inversions, improving detection rates and understanding their role in diseases, with high resolution and specificity, enabling the identification of small inversions and providing a mechanism for precise inversion detection.
Implementation Method 1
hybridizing the probes to the sister chromatids; and detecting the hybridized probes
Implementation Method 2
labeled probes to one chromatid and not the other using fluorescence microscopy
Data Source
AI summary
A kit for the characterization of chromosomal inversions using single-stranded probes that are either all identical or all complementary to a single-stranded chromatid is described. Reporter species are attached to oligonucleotide strands designed such that they may hybridize to portions of only one of a pair of single-stranded sister chromatids which may be prepared by the CO-FISH procedure. If an inversion has occurred, these marker probes will be detected on the second sister chromatid at the same location as the inversion on the first chromatid. The kit includes non-repetitive probes that are either all identical or all complementary to at least a portion of a target DNA sequence of only one DNA strand of only one chromatid and may in some embodiments include reagents suitable for performing CO-FISH and/or reagents for hybridizing the probes to the target DNA sequence.