Partition Indexing Oligonucleotides for Single-Cell Spatial Mapping
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Solution Overview
Problem
Current methods for determining gene expression in single cells face challenges such as doublet formation, noise in sequencing data, and the need to associate sequencing data with phenotypic information, leading to inaccurate interpretation of expression profiles and noise in results.
Innovation Solution
The method involves using partition indexing oligonucleotides with unique sequences to assign sequencing data to specific partitions, identify noise partitions, and associate sequencing data with phenotypic data by barcoding nucleic acid targets and obtaining sequencing data from these partitions, allowing for accurate assignment and reduction of noise.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If cells are co-localized with barcoded reagent beads in compartments to enable massively parallel gene expression measurement, then measurement throughput is improved, but doublet formation increases causing inaccurate expression profiles
Solution Approach 1:
The invention segments the barcode space by introducing partition indexing sequences that divide the sequencing data into distinct partitions. Each partition corresponds to a specific cell or compartment, allowing the system to distinguish between single cells and doublets by analyzing which partition the sequencing reads map to. This segmentation enables accurate single-cell resolution even when cells are co-localized with barcoded beads.
Solution Approach 2:
The partition indexing oligonucleotides serve as an intermediary element between the barcoded reagent beads and the sequencing reads. These indexing sequences are attached to the beads and provide a unique identifier for each compartment, mediating the connection between physical co-localization and digital identification in the sequencing data.
2Measurement precision
If partition indexing oligonucleotides with unique sequences are used to assign sequencing data to partitions, then data assignment precision is improved, but device complexity increases
Solution Approach 1:
The partition indexing oligonucleotides perform multiple functions: they serve as compartment identifiers, enable data assignment to partitions, and provide a basis for distinguishing single cells from doublets. By making this single element multi-functional, the system achieves high precision without proportionally increasing complexity.
Solution Approach 2:
The invention changes the parameter of sequence uniqueness from the barcode level to the partition indexing level. Instead of requiring entirely unique barcodes for each cell, the system uses unique partition indexing sequences at the compartment level, which simplifies the overall system while maintaining precision through the combination of partition indexing and barcode reading.
3Productivity
If sequencing data from multiple partitions is pooled together, then productivity is improved, but noise in sequencing data increases making it difficult to associate phenotypic information
Solution Approach 1:
The invention segments the sequencing data into distinct partitions based on the partition indexing sequences, allowing each partition to be processed and analyzed separately. This segmentation maintains data quality by preventing mixing of signals from different cells or compartments, while still enabling efficient pooled processing through standardized workflows that handle multiple partitions simultaneously.
Solution Approach 2:
The partition indexing sequences provide a feedback mechanism that allows the system to track and verify the origin of each sequencing read. By knowing which partition each read comes from, the system can filter out noise and misassignments, ensuring that phenotypic information can be reliably associated with the correct cell or compartment.
Data Source
AI summary
Disclosed herein include systems, methods, compositions, and kits for methods of assigning sequencing data to partitions. There are provided, in some embodiments, methods of associating sequencing data and phenotypic data of single cells. There are provided, in some embodiments, methods of reducing noise in sequencing data. Disclosed herein include partition indexing oligonucleotides comprising a partition indexing sequence. The partition indexing oligonucleotides can be associated with partitions. Partition indexing oligonucleotides situated within the same partition can comprise the same partition indexing sequence. Partition indexing oligonucleotides situated within different partitions comprise different partition indexing sequences.