Single-Cell Nucleic Acid Barcoding for Minority Population Attribution
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Solution Overview
Problem
Current nucleic acid sequencing technologies struggle to effectively analyze and characterize sub-populations of cells, particularly those representing a minority within a biological sample, due to biases in amplification and attribution methods, leading to inaccurate data and inability to identify individual cell contributions.
Innovation Solution
The method involves compartmentalizing nucleic acids from individual cells into discrete partitions, attaching unique barcode sequences, and sequencing these to attribute characteristics back to the individual cells, allowing for precise characterization of nucleic acids from small populations or individual cells without amplification biases.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional nucleic acid sequencing methods are used, then sequencing can be performed on bulk cell populations, but amplification biases prevent accurate characterization of sub-populations and minority cell contributions
Solution Approach 1:
The patent segments bulk cell populations into individual cells or small groups through single-cell partitioning techniques. Each partition contains nucleic acids from a single cell or small group of cells, allowing independent sequencing and attribution. This segmentation eliminates amplification biases that occur in bulk methods, as each partition is processed separately without cross-contamination or dominant signal interference from majority cell types.
Solution Approach 2:
The patent introduces barcode sequences as intermediary markers that are attached to nucleic acids from individual cells. These barcodes serve as mediators between the physical cell and the sequencing data, enabling accurate attribution of sequencing results to specific cells. The barcodes allow differentiation between cells that would otherwise be indistinguishable in bulk analysis, resolving the attribution accuracy problem.
2Quantity of substance
If amplification methods are used to increase sequencing signal, then sequencing sensitivity improves, but amplification biases distort the representation of minority cell populations
Solution Approach 1:
The patent performs preliminary actions by attaching barcode sequences to nucleic acids from individual cells before amplification. This preliminary barcoding ensures that even low-abundance sequences from minority cells are tagged and identifiable before amplification occurs. The barcodes are attached at the single-cell level, so amplification of individual partitions does not create biases, as each partition's barcode-coded sequences are independently amplified and tracked.
Solution Approach 2:
The patent uses barcode sequences as copies or identifiers that travel with the nucleic acids through the sequencing process. Instead of relying on the quantity of nucleic acid molecules for identification, the barcodes provide a discrete, copyable identifier that maintains information about the original cell source. This copying mechanism allows accurate tracking of minority cell contributions even when nucleic acid amounts are low.
3Productivity
If bulk sequencing is performed on heterogeneous samples, then throughput is maintained, but individual cell contributions cannot be identified
Solution Approach 1:
The patent segments the heterogeneous sample into individual cell partitions while maintaining high throughput through parallel processing. Multiple partitions containing single cells or small groups of cells can be sequenced simultaneously using high-throughput sequencing platforms. The segmentation is achieved through microfluidic devices or droplet-based systems that can process many partitions in parallel, thus maintaining productivity while preserving cell-level information through the barcode attachment method.
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.