Systems and methods to detect rare mutations and copy number variation

What is this patent about?

’376 is related to the field of genetic analysis and molecular diagnostics, specifically focusing on the detection of rare mutations and copy number variations within cell-free polynucleotides. The technology addresses the inherent challenges of identifying low-frequency genetic signals, such as those derived from tumors or fetuses, which are often obscured by the high background noise and distortion typical of standard next-generation sequencing workflows.

The underlying idea behind ’376 is to utilize a digital sequencing approach that employs molecular tagging to distinguish true genetic variants from artifacts introduced during laboratory processing. By assigning identifiers to individual parent molecules before amplification, the system can collapse multiple sequencing reads into high-fidelity consensus sequences, effectively filtering out errors and providing a more accurate quantitative measure of the original genetic material.

The claims of ’376 focus on a method for preparing DNA libraries for sequencing by attaching a specific number of molecular barcodes to a population of DNA molecules. The number of unique barcodes, denoted as n, is strategically calibrated to be at least 2 but no more than 100,000 times the mean number of expected duplicate molecules that share identical start and stop positions, ensuring that cognate fragments can be uniquely identified even in samples containing up to 300,000 haploid human genome equivalents.

In practice, the invention works by ligating these molecular barcodes to cell-free DNA fragments, followed by amplification and selective enrichment of specific genomic regions of interest, such as actionable oncogenes. This targeted approach allows for the deep sequencing of progeny polynucleotides, which are then grouped into families based on their barcodes and genomic coordinates to reconstruct the sequence of the original parent molecule with high sensitivity.

This methodology differs from prior approaches by optimizing the conversion efficiency of the library preparation and using a mathematically defined diversity of tags to handle the natural overlap of fragmented DNA. By focusing on the relationship between the number of tags and the expected number of duplicate molecules, the system achieves a sensitivity threshold as low as 0.1%, enabling the detection of rare variants that would otherwise be lost in the baseline noise of conventional analog sequencing.