Barcoded Bead Partitioning for Single-Cell Sequencing
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Solution Overview
Problem
Current nucleic acid sequencing technologies are ineffective at identifying and characterizing cells at the single cell level, often averaging genetic material from cell populations and failing to accurately represent minority cell constituents, leading to inaccurate data, especially in heterogeneous samples.
Innovation Solution
A method and system that compartmentalize individual cells or small populations into discrete partitions, using labelling agents with nucleic acid barcode sequences to attribute characteristics back to specific cells, allowing for precise characterization of cell surface features and nucleic acids.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If nucleic acid sequencing is performed on cell populations using conventional methods, then sequencing can be conducted, but the results average genetic material and fail to accurately represent minority cell constituents
Solution Approach 1:
The patent segments the cell population into individual cells by partitioning them into separate droplets or wells. Each cell is isolated in its own partition, allowing independent processing and characterization. This segmentation enables the preservation of individual cell information, including minority cell constituents, rather than averaging across the entire population.
Solution Approach 2:
The patent introduces barcoded beads as intermediaries that carry unique molecular identifiers (UMIs) and capture oligonucleotides. These beads mediate the connection between individual cells and the sequencing process, enabling tracking and attribution of genetic material back to specific cells. The barcoded beads serve as carriers that preserve cell-specific information through the processing workflow.
2Measurement precision
If conventional nucleic acid sequencing methods are used, then the process is simpler, but it cannot identify or attribute characteristics to specific individual cells
Solution Approach 1:
The patent performs preliminary actions by pre-loading beads with barcodes and capture oligonucleotides before cell partitioning. The barcoding system is prepared in advance, and cells are subsequently assigned to partitions containing these pre-prepared beads. This preliminary preparation simplifies the overall process by establishing the identification framework before the complex partitioning and sequencing steps.
Solution Approach 2:
The patent uses barcoded beads as copies or proxies for individual cells. Each bead carries a unique barcode that serves as a digital copy or identifier for the cell it captures. This copying mechanism enables the tracking and identification of individual cells through their barcode representations, allowing complex cell population analysis to be managed through simpler bead-based proxies.
3Reliability
If ensemble approaches are used for nucleic acid sequencing, then processing is more straightforward, but accurate data on individual cells and minority cell contributions cannot be obtained
Solution Approach 1:
The patent segments the heterogeneous cell population into individual partitions, ensuring that each cell's genetic material is processed separately. This segmentation prevents the averaging effect that occurs in ensemble approaches and preserves the unique characteristics of each cell, including rare minority cell constituents. The segmentation is achieved through droplet generation or well-based partitioning systems.
Solution Approach 2:
The patent implements feedback through the barcoding system, where unique molecular identifiers (UMIs) on beads provide information that feeds back into the data analysis process. The barcodes enable tracking of which cell each genetic read originated from, allowing for accurate attribution and quantitative analysis. This feedback mechanism ensures reliable data by maintaining the connection between sequencing reads and their source cells throughout the process.
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
Enables accurate characterization of individual cells or small populations, overcoming the limitations of ensemble approaches by providing precise data on cell surface features and nucleic acids, even in heterogeneous samples, and allowing for the identification of minority cell contributions.
Implementation Method 1
the at least one labelling agent is (i) capable of binding to a cell surface feature of the cell
Implementation Method 2
subjecting the nucleic acid molecule to sequencing to identify the labelling agent or the cell
Data Source
AI summary
The present disclosure provides compositions, methods, systems, and devices for polynucleotide processing and analyte characterization. Such polynucleotide processing may be useful for a variety of applications, including analyte characterization by polynucleotide sequencing. The compositions, methods, systems, and devices disclosed herein generally describe barcoded oligonucleotides, which can be bound to a bead, such as a gel bead, useful for characterizing one or more analytes including, for example, protein (e.g., cell surface or intracellular proteins), genomic DNA, and RNA (e.g., mRNA or CRISPR guide RNAs). Also described herein, are barcoded labelling agents and oligonucleotide molecules useful for “tagging” analytes for characterization.