Barcode-Tagged Nucleic Acid Partitioning for Minority Cell Sequencing
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Solution Overview
Problem
Current nucleic acid sequencing technologies struggle to effectively identify and characterize sub-populations of cells in biological samples, particularly those representing a minority of the overall sample, due to ensemble-based methods that are biased towards majority constituents and fail to preserve the ratios of minority and majority components during geometric amplification.
Innovation Solution
The method involves compartmentalizing nucleic acids from individual cells or small populations into discrete partitions, where unique barcodes are attached to the nucleic acids, allowing for their characterization and attribution back to the individual cells, even in heterogeneous mixtures, using techniques such as reverse transcription and barcode-based sequencing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If ensemble-based sequencing methods are used, then high-throughput analysis is achieved, but the ability to identify and characterize sub-populations of cells is lost
Solution Approach 1:
The patent divides the cell population into individual cells or small groups, each assigned a unique barcode. This segmentation allows high-throughput sequencing to be performed on individual units while maintaining the ability to identify and characterize sub-populations through their unique barcodes, resolving the contradiction between throughput and sub-population identification.
Solution Approach 2:
The patent introduces barcode sequences as intermediary molecules that link individual cells to sequencing data. These barcodes serve as mediators that enable both high-throughput processing and precise identification of sub-populations, as each barcode can be uniquely tracked through the sequencing process.
2Quantity of substance
If geometric amplification is used, then nucleic acid signal is enhanced, but the ratios of minority and majority components are not preserved
Solution Approach 1:
By segmenting the sample into individual cells or small groups with unique barcodes, the patent enables amplification to occur within isolated units. This prevents the amplification bias that occurs in ensemble methods, allowing the original ratios of minority and majority components to be preserved while still achieving sufficient signal through localized geometric amplification.
3Measurement precision
If single-cell analysis is performed, then accurate characterization of individual cells is achieved, but the throughput is reduced
Solution Approach 1:
The patent merges the advantages of single-cell analysis with high-throughput processing by using barcode sequences that can be read through pooled samples. Individual cells are analyzed with the precision of single-cell methods, while the barcode-based pooling enables high-throughput characterization of many cells simultaneously, resolving the contradiction between precision and productivity.
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 enables the accurate characterization of individual cells or small populations within larger samples, preserving the ratios of minority and majority components, and providing high-throughput analysis of low nucleic acid inputs, overcoming the limitations of existing ensemble sequencing methods.
Implementation Method 1
attaching, to the nucleic acids, oligonucleotides that comprise a common nucleic acid barcode sequence
Implementation Method 2
using techniques such as reverse transcription and barcode-based sequencing
Data Source
AI summary
The present disclosure provides compositions, methods, systems, and devices for polynucleotide processing. Such polynucleotide processing may be useful for a variety of applications, including polynucleotide sequencing.