Single-Cell Nucleic Acid Barcoding With Split Oligonucleotide Tags
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Solution Overview
Problem
Current methods for barcoding nucleic acids in single-cell analysis are costly and time-consuming due to the need for high concentrations of barcode molecules, often requiring bead-based approaches that are expensive and error-prone.
Innovation Solution
A method involving the co-partitioning of single cells with single copies of oligonucleotides having different barcode sequences, where the first oligonucleotides are attached to target nucleic acids, followed by amplification to generate copies with combined barcode sequences, reducing the need for excess second oligonucleotides.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If high concentrations of barcode molecules are used for barcoding, then barcoding reliability is improved, but material cost and expense increase
Solution Approach 1:
The barcode is divided into two separate oligonucleotide components (first and second oligonucleotides) that are co-partitioned with the cell. Each oligonucleotide contains a distinct barcode sequence, and their combination provides the full barcode information. This segmentation allows each component to be present at lower concentrations while maintaining overall barcoding reliability through their combined presence.
Solution Approach 2:
The invention transitions from using a single high-concentration barcode oligonucleotide to using two lower-concentration oligonucleotides that combine their barcode sequences. This dimensional change in the barcoding approach (from one component to two components) reduces the quantity requirement of each individual oligonucleotide while preserving barcoding functionality.
2Reliability
If bead-based methods are used for barcoding, then barcoding capability is improved, but manufacturing complexity and time increase
Solution Approach 1:
The invention extracts the barcode functionality from the complex bead structure and implements it through simple free-floating oligonucleotides. By removing the bead carrier, the method eliminates the complexities of bead synthesis, handling, and error-prone procedures while retaining the essential barcoding capability through the oligonucleotide sequences alone.
Solution Approach 2:
The invention replaces expensive, complex beads with simple, inexpensive oligonucleotides that can be synthesized through standard PCR amplification. These oligonucleotides serve as disposable barcoding elements that are easy to manufacture and do not require the complex infrastructure needed for bead production.
3Reliability
If multiple copies of barcode oligonucleotides are provided, then barcoding completeness is improved, but cost intensity increases
Solution Approach 1:
The invention merges the barcoding function into two separate oligonucleotide components that are co-partitioned with the cell. By combining their individual barcode sequences, they achieve complete barcoding coverage without requiring excessive amounts of each individual oligonucleotide, thereby reducing cost intensity while maintaining barcoding completeness.
Data Source
AI summary
The current invention provides a cost effective and reliable method for barcoding of target nucleic acids from single cells.