Functionalized Gel Beads for Barcode-Encoded Analyte Identification
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Solution Overview
Problem
Current methods for processing biological samples in partitioned environments, such as PCR and sequencing, lack efficient means for controlled analysis and processing of analytes like nucleic acid molecules and proteins, particularly in generating and utilizing barcode molecules for accurate identification and quantification.
Innovation Solution
The method involves generating barcode molecules by combinatorially assembling molecules on supports like beads, using split pool ligation reactions, to create distinct barcode sequences for controlled analysis and processing of analytes, allowing for precise identification and quantification.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional methods (PCR, sequencing) are used for processing biological samples, then sample processing can be performed, but efficient controlled analysis and processing of analytes with accurate identification and quantification is lacking
Solution Approach 1:
The sample processing system is segmented into multiple partitions (droplets or wells), with each partition containing specific analytes and corresponding barcode molecules. This segmentation enables parallel processing of multiple analytes simultaneously, improving productivity while maintaining measurement precision through isolated, controlled environments for each analyte-barcode pair.
Solution Approach 2:
The invention uses fluorescently labeled barcode molecules that emit detectable signals when bound to target analytes. Each barcode can be distinguished by its fluorescent properties, enabling accurate identification and quantification of multiple analytes simultaneously, thus improving measurement precision without sacrificing processing efficiency.
2Measurement precision
If barcode molecules are generated for each analyte to improve identification accuracy, then measurement precision improves, but the complexity of generating and managing multiple barcode types increases
Solution Approach 1:
Multiple barcode molecules with different sequences are combined on a single bead support structure. This merging approach allows multiple analytes to be identified using barcodes from the same physical support, reducing the number of separate components needed and simplifying the overall system while maintaining the ability to distinguish between different analytes through sequence-specific hybridization.
Solution Approach 2:
The bead support structure is designed to universally hold multiple types of barcode molecules, making it a multi-functional platform that can identify various analytes. This universal support reduces device complexity by eliminating the need for separate supports for each barcode type, while still enabling precise identification through sequence-specific binding.
3Measurement precision
If multiple barcode molecules are attached to each bead to improve analyte identification, then measurement precision improves, but the manufacturing process becomes more complex
Solution Approach 1:
Barcode molecules are pre-synthesized and attached to bead supports in a controlled manufacturing process before use. This preliminary action allows for standardized production of multi-barcode beads, simplifying the manufacturing process by separating barcode attachment from the final assay procedure, and enabling quality control during production rather than during use.
Solution Approach 2:
The manufacturing process utilizes controlled parameters such as hybridization temperature, salt concentration, and incubation time to optimize barcode attachment to beads. By adjusting these parameters, the process achieves high precision barcode-bead binding while maintaining ease of manufacture through standardized, reproducible conditions that can be scaled.
Data Source
AI summary
The present disclosure provides methods of generating supports (e.g., beads) comprising barcode molecules coupled thereto. A barcode molecule coupled to a support may comprise a barcode sequence and a functional sequence. A barcode molecule may be generated using two or more ligation reactions in a combinatorial fashion. A support comprising two or more different barcode molecules may be useful for analyzing or processing one or more analytes such as nucleic acid molecules, proteins, and/or perturbation agents.