DNA-Barcoded Bead Libraries for Single-Cell Picowell Screening
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Solution Overview
Problem
Current high-throughput drug screening methods face limitations in efficiently partitioning different drugs to different cells, particularly in large-scale assays with over 100,000 wells, and lack efficient methods for single-bead dispensing in such setups, leading to bottlenecks in RNA profiling and drug examination capacity.
Innovation Solution
A system utilizing picowell arrays with beads containing multiple copies of compounds and DNA tags for high-throughput screening, where each bead has DNA barcodes encoding compound identity, allowing for controlled release and biological activity screening, and incorporating caps to prevent evaporation and leakage, with methods for creating concatenated or orthogonal DNA barcodes and using cleavable linkers for compound delivery.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional high-throughput screening methods are used with microwell or nanowell arrays, then screening capacity is limited to manageable scales, but throughput and drug examination capacity become bottlenecks when scaling to over 100,000 wells
Solution Approach 1:
The system partitions compounds and cells into picowell compartments, with each picowell containing a single bead and single cell. This segmentation enables independent high-throughput screening of millions of compound-cell interactions simultaneously, resolving the throughput bottleneck while maintaining manageable individual unit complexity
Solution Approach 2:
Beads serve as intermediaries that carry both compounds and DNA barcodes, enabling simultaneous delivery of chemical perturbations and identity tracking to picowells. This intermediary approach allows complex information (compound identity, synthesis history) to be transmitted without increasing system complexity
2Measurement precision
If DNA barcodes are used to encode compound identity on beads, then compound identification precision is improved, but manufacturing and sequencing complexity increases
Solution Approach 1:
DNA barcodes encoding compound identity and synthesis history are incorporated onto beads during the compound synthesis process itself, rather than being added separately later. This preliminary action ensures precise tracking from the moment of compound creation while streamlining manufacturing by combining synthesis and tagging operations
Solution Approach 2:
Each bead carries multiple copies of the same DNA barcode sequence, ensuring robust signal for sequencing detection. This copying approach improves identification precision by providing sufficient template material while maintaining manufacturing simplicity through standardized barcode incorporation protocols
3Reliability
If beads contain multiple copies of compounds for screening, then screening reliability is improved, but compound release control becomes more difficult
Solution Approach 1:
The system employs cleavable linkers with specific chemical parameters that allow controlled release of compounds from beads under defined conditions (e.g., specific pH, enzymatic treatment, or chemical triggers). This parameter-based control enables reliable compound delivery during screening while facilitating precise release when needed, without compromising ease of operation
4Measurement precision
If picowell arrays are used for single-cell screening, then measurement precision of cellular responses is improved, but device complexity and handling difficulty increase
Solution Approach 1:
The system combines multiple functions into the bead component: compound delivery, identity encoding via DNA barcodes, and perturbation application. This merging reduces the number of separate operations needed in picowells, thereby decreasing device complexity and handling difficulty while maintaining single-cell measurement precision
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 high-throughput screening of millions to billions of compounds with precise identification and controlled release, overcoming throughput limitations and enabling efficient drug discovery and diagnostics by facilitating single-cell perturbation and response analysis.
Implementation Method 1
the bead-bound compound is coupled to the bead by way of a cleavable linker, and cleaving the cleavable linker releases the bead-bound compound from the bead to generate a released compound
Implementation Method 2
uses a splint oligonucleotide (splint oligo) that is capable of hybridizing to a partially made bead-bound DNA barcode
Data Source
AI summary
This application provides a bead with a covalently attached chemical compound and a covalently attached DNA barcode and methods for using such beads. The bead has many substantially identical copies of the chemical compound and many substantially identical copies of the DNA barcode. The compound consists of one or more chemical monomers, where the DNA barcode takes the form of barcode modules, where each module corresponds to and allows identification of a corresponding chemical monomer. The nucleic acid barcode can have a concatenated structure or an orthogonal structure. Provided are a method for sequencing the bead-bound nucleic acid barcode, for cleaving the compound from the bead, and for assessing biological activity of the released compound.