Modified Reference Genome for SMA Paralog Differentiation
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Solution Overview
Problem
Current methods for diagnosing spinal muscular atrophy (SMA) based on whole genome sequencing data face challenges in accurately distinguishing between SMN1 and SMN2 genes due to their high similarity, leading to ambiguous read alignments and difficulties in determining affected or carrier status.
Innovation Solution
The method involves aligning whole genome sequencing reads to a modified reference genome sequence with SMN2 converted to a string of Ns, allowing for the counting of quasi-alleles and adjustment for coverage to determine the number of functional SMN1 gene copies, and using graph-based methods to distinguish between SMN1 and SMN2 genes.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If whole genome sequencing reads are aligned to a standard reference genome containing both SMN1 and SMN2 genes, then the alignment process captures all genomic information, but the high similarity between SMN1 and SMN2 causes ambiguous read alignments and inaccurate diagnosis
Solution Approach 1:
The patent extracts the SMN2 gene sequence from the reference genome, replacing it with Ns (ambiguous bases). This creates a modified reference genome that contains only SMN1 sequences, eliminating the ambiguity caused by SMN1-SMN2 similarity while preserving all SMN1 diagnostic information
Solution Approach 2:
The patent segments the analysis into two distinct parts: (1) alignment to the modified reference genome (SMN2-depleted) to accurately count SMN1 copies, and (2) separate analysis to determine SMN2 copy number. This segmentation resolves the alignment ambiguity by handling each gene separately
2Productivity
If standard alignment methods are used without modification, then the process is simple and fast, but the high similarity between SMN1 and SMN2 genes leads to read misalignment and false results
Solution Approach 1:
The patent performs preliminary modification of the reference genome by replacing SMN2 sequences with Ns before the alignment process. This preliminary action prevents read misalignment from occurring in the first place, maintaining fast processing speeds while ensuring reliable results
3Quantity of substance
If the reference genome includes both SMN1 and SMN2 sequences, then comprehensive genomic coverage is achieved, but distinguishing between the two genes becomes difficult due to their near-identical sequences
Solution Approach 1:
The patent introduces an intermediary modified reference genome that acts as a mediator between the raw sequencing reads and the final diagnosis. By replacing SMN2 sequences with Ns in this intermediary reference, the system enables clear differentiation of SMN1 reads while preserving comprehensive genomic coverage for other regions
Data Source
AI summary
Disclosed herein are systems and methods for spinal muscular atrophy (SMA) diagnosis from whole genome sequencing data. In one embodiment, a method comprises aligning whole genome sequencing (WGS) reads of a subject's sample to a modified reference sequence such as a modified reference genome sequence. After counting the reads supporting quasi-alleles at select positions of the reference sequence, the method can adjust for coverage and determine a number of functional SMNl gene copies. The method can determine affected or carrier status of the subject based on the copy number of functional SMNl gene copies.