Nucleic Acid Sequencing Phasing via Intramolecular Barcode Distribution
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Next Generation Sequencing (NGS) technologies face limitations in determining whether genetic variations originate from the same or different nucleic acid molecules due to limitations in sequencing read lengths, making it challenging to phase genetic variations accurately.
Innovation Solution
A method involving the use of nucleic acid strands with adaptors containing elongation sequences and molecular barcodes, where the elongation sequence is complementary to a portion of the nucleic acid sequence, allowing for intramolecular elongation and generation of stem-loop structures, enabling the distribution of barcodes throughout the nucleic acid molecule for improved sequencing and phasing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If standard NGS sequencing is used, then high-throughput sequencing can be achieved, but sequencing read lengths are limited making it difficult to determine molecular origin of genetic variations
Solution Approach 1:
The adaptor is segmented into distinct functional regions: a molecular barcode region for identification and an elongation sequence region for template-directed synthesis. This segmentation allows the barcode to be distributed to multiple locations along the nucleic acid molecule through intramolecular elongation, enabling molecular origin tracking while maintaining high sequencing throughput
Solution Approach 2:
The adaptor serves as an intermediary molecule that bridges the template nucleic acid and the sequencing process. It carries the molecular barcode and provides the elongation sequence that mediates the intramolecular synthesis, thereby linking the original molecular identity to the sequenced fragments without compromising sequencing capacity
2Measurement precision
If adaptors with elongation sequences are used for intramolecular elongation, then barcodes can be distributed throughout the nucleic acid molecule, but the process complexity increases
Solution Approach 1:
The adaptor design enables self-service through intramolecular elongation where the elongation sequence within the same adaptor molecule serves as the template for synthesizing new copies of the barcode. This self-directed process distributes barcodes throughout the nucleic acid molecule without requiring external intervention or complex additional reagents, reducing overall process complexity despite the enhanced functionality
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 enhances the ability to phase genetic variations by evenly distributing barcodes along the nucleic acid sequence, reducing coverage bias and allowing for accurate determination of the molecular origin of genetic variations, thereby improving the interpretation of genetic information.
Implementation Method 1
annealing said elongation sequence to said portion of said nucleic acid sequence in said nucleic acid strand, thereby generating a partially-duplexed nucleic acid strand
Implementation Method 2
extending said elongation sequence with a polymerase using said 5′ portion of said partially-duplexed nucleic acid strand as a template, thereby generating an extended nucleic acid
Data Source
AI summary
The present disclosure provides methods and compositions for molecular tagging of complex populations of nucleic acid molecules. The disclosure provides methods and compositions to obtain phase information of tagged nucleic acid molecules from high-throughput nucleic acid sequencing data.