Barcoded Cell Partitioning for Individual Polynucleotide Characterization
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Solution Overview
Problem
Existing nucleic acid sequencing technologies struggle to accurately characterize and attribute genomic and proteomic information to individual cells or small populations of cells, often leading to inaccurate data due to ensemble processing methods that favor majority constituents and fail to preserve the characteristics of minority cells.
Innovation Solution
A method involving the compartmentalization of individual cells or small cell populations into discrete partitions, where each partition is labeled with unique barcodes to attribute cell surface features and nucleic acids back to the individual cells, using labelling agents and oligonucleotides to facilitate sequencing and characterization.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If ensemble processing methods are used for nucleic acid sequencing, then throughput and productivity are improved, but measurement precision and reliability of individual cell characterization deteriorate
Solution Approach 1:
The patent partitions a population of cells into individual discrete partitions (droplets or wells), with each partition containing at most one cell. This segmentation allows simultaneous processing of many individual cells through high-throughput sequencing while preserving the ability to attribute sequencing reads to specific individual cells via cell barcodes, thereby resolving the contradiction between throughput and individual cell characterization accuracy.
2Productivity
If ensemble processing methods are used, then productivity is improved, but loss of information about minority cell populations increases
Solution Approach 1:
By segmenting the cell population into individual partitions and tagging each cell with a unique cell barcode, the patent enables simultaneous high-throughput processing and preservation of individual cell information. This allows rare minority cell populations to be identified and characterized without being obscured by majority constituents, eliminating information loss while maintaining productivity.
Solution Approach 2:
The patent introduces cell barcodes as intermediary molecules that link individual cells to their sequencing reads. Each cell is tagged with a unique cell barcode that is incorporated into the sequencing library, allowing computational attribution of reads to specific cells. This intermediary enables the preservation of individual cell information through ensemble processing, preventing loss of minority cell characteristics.
3Measurement precision
If partitioning into discrete compartments is implemented, then measurement precision of individual cells is improved, but device complexity increases
Solution Approach 1:
The patent employs microfluidic hydrodynamic focusing and droplet generation techniques to automatically partition cells into discrete compartments. This hydraulic approach enables high-throughput partitioning without complex mechanical manipulation, reducing device complexity while achieving precise individual cell isolation and characterization.
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 and attribution of genomic and proteomic information to individual cells, overcoming biases in ensemble processing and preserving the unique characteristics of minority cell populations.
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
an anchor oligonucleotide that is capable of interacting with the reporter oligonucleotide barcode
Implementation Method 3
in the partition, synthesizing a nucleic acid molecule comprising at least a portion of the nucleic acid barcode sequence
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.