Single Cell Nucleic Acid Processing via Sequential Co-Partitioning
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Solution Overview
Problem
Current methods for processing biological samples often face challenges in simultaneously analyzing multiple types of biomolecules, such as RNA and DNA, from a single cell without altering or destroying one type of biomolecule.
Innovation Solution
A method involving the co-partitioning of a cell bead and a barcode bead in a partition, where the cell bead contains both RNA and DNA, and the barcode beads have nucleic acid barcode sequences. The method applies specific conditions to couple target nucleic acid molecules with barcode molecules, generating barcoded nucleic acid molecules that can be sequenced to associate the molecules with their origin in the cell.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional methods are used to process multiple types of biomolecules from a single cell, then one type of biomolecule can be obtained, but another type of biomolecule is altered or destroyed
Solution Approach 1:
The processing method is divided into sequential stages: first processing one type of nucleic acid molecule (e.g., RNA) while preserving another type (e.g., DNA), then subsequently processing the second type. This temporal segmentation allows each biomolecule type to be analyzed under optimized conditions without interfering with the other, thus maintaining integrity while achieving versatility.
Solution Approach 2:
The method performs preliminary actions to capture and preserve one type of biomolecule (e.g., RNA capture using oligo-dT beads) before processing another type (e.g., DNA extraction). This preliminary capture ensures that the first biomolecule type is secured and preserved in a stable form, preventing alteration or destruction during subsequent processing steps for the second type.
2Reliability
If sequential processing steps are implemented to preserve multiple biomolecule types, then integrity is maintained, but processing time increases
Solution Approach 1:
The method merges multiple processing objectives into a unified workflow where capture beads and processing reagents are combined in the same reaction environment. For example, oligo-dT beads for RNA capture and DNA extraction reagents are present together in the partition, allowing sequential processing of different biomolecule types without requiring separate physical processing steps, thus reducing overall processing time while maintaining integrity.
3Measurement precision
If multiple processing conditions are applied to different biomolecule types, then accurate analysis is achieved, but system complexity increases
Solution Approach 1:
The patent uses intermediary capture beads (e.g., oligo-dT beads, magnetic beads) that mediate between different processing conditions for different biomolecule types. These beads serve as universal platforms that can facilitate RNA capture under one set of conditions and subsequent DNA processing under different conditions, simplifying the overall system by providing a common interface for handling multiple biomolecule types with different processing requirements.
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 method allows for the simultaneous analysis and identification of multiple types of nucleic acid molecules from a single cell, preserving the integrity of each type of biomolecule and enabling accurate association of sequencing reads with their cellular origin.
Implementation Method 1
a first target nucleic acid molecule of the first type to couple to a first barcode molecule of the first set of barcode molecules to generate a first barcoded nucleic acid molecule
Implementation Method 2
a second target nucleic acid molecule of the second type to couple to a third barcode molecule of the second set of barcode molecules to generate a second barcoded nucleic acid molecule
Implementation Method 3
a fourth barcode molecule of the second set of barcode molecules to couple to the second barcode molecule to generate a composite barcode molecule
Data Source
AI summary
Provided herein are methods, compositions, and systems for multiplexed analysis of individual cells or cell populations. Cells encapsulated in beads and/or biomolecules are sequentially co-partitioned, allowing for analysis of two different types of biomolecules (e.g., RNA and DNA). The present invention leverages different polymer dissociation mechanisms, accompanied with barcoding of biomolecules (e.g., nucleic acid molecules) for multiplexed measurements in single cells. Sequential co-partitioning and barcode technology enables identification and quantitation of DNA and RNA from single cells.