Polynucleotide Barcode Insertion for Genome Sequencing Alignment
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Solution Overview
Problem
Next-generation sequencing (NGS) technologies face challenges in accurately aligning DNA sequence reads due to errors and complexities, leading to incorrect variant detection and increased costs in sequencing entire genomes.
Innovation Solution
The method involves inserting polynucleotide barcodes into a target DNA or RNA sequence, sequencing multiple times, and determining the sequence by stitching together reads with overlapping barcodes to improve alignment and variant detection accuracy.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If NGS methods use computational techniques to align read data, then sequencing throughput increases, but alignment accuracy deteriorates due to errors and complexities
Solution Approach 1:
The patent introduces polynucleotide barcodes as intermediary markers inserted at known locations within the target polynucleotide. These barcodes serve as reference points that facilitate accurate alignment of sequence reads by providing identifiable landmarks, thereby resolving the contradiction between high-throughput sequencing and alignment accuracy.
2Reliability
If multiple sequence reads are generated in parallel, then sequencing cost decreases, but variant detection accuracy deteriorates due to misalignment
Solution Approach 1:
Polynucleotide barcodes are inserted at known locations to serve as reference markers. These barcodes enable accurate alignment of multiple parallel sequence reads by providing identifiable landmarks, thereby maintaining variant detection accuracy while preserving the cost efficiency of parallel sequencing.
3Loss of time
If read data is aligned using conventional methods, then processing time is reduced, but sequence determination accuracy deteriorates in repeat regions
Solution Approach 1:
Polynucleotide barcodes are inserted at known locations within the target polynucleotide before sequencing. These pre-placed barcodes serve as reference markers that facilitate rapid and accurate alignment, particularly in repeat regions, by providing known positional information that resolves ambiguities without requiring extensive computational processing.
4Measurement precision
If polynucleotide barcodes are inserted at multiple locations, then alignment accuracy improves, but device complexity increases
Solution Approach 1:
The target polynucleotide is divided into multiple segments with polynucleotide barcodes inserted at specific locations within each segment. This segmentation approach provides multiple reference points for alignment, improving accuracy while maintaining manageable process complexity through systematic placement of standardized barcode elements.
Data Source
AI summary
Contemporary gene sequencing techniques, including “Next Generation Sequencing” techniques, can include sequencing a plurality of fragments of a target polynucleotide. These fragment sequences are then used to determine a sequence for the target as a whole. This can include aligning the fragment sequences to each other anchor to a reference genome. However, the limitations of existing sequencing techniques, and the often repetitive or otherwise difficult-to-sequence structure of natural polynucleotides, means that it can be difficult and/or expensive to generate accurate sequences. Methods provided herein include inserting polynucleotide ‘barcodes’ into a target polynucleotide prior to fragmentation or other sequencing processes. These inserted barcodes can improve the accuracy of sequences generated for the target by adding ‘noise’ into the target, allowing subsequent sequencing techniques (e.g., alignment, stitching, etc.) to more accurately estimate the target-plus-barcodes sequence. The barcodes can then be removed to provide the sequence of the target polynucleotide.