Automated Cell Isolation for Genome Editing Efficiency
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Solution Overview
Problem
Current nucleic acid-guided nuclease gene editing methods face challenges in efficiently identifying and enriching edited cells due to rapid depletion of edited cell types and selective enrichment of unedited cells in bulk or multiplex formats, where only a small fraction of cells survive double-strand DNA breaks.
Innovation Solution
The development of instruments and methods that enable automated high-throughput screening and substantial isolation of cells, allowing for clonal growth, normalization, and cherry picking of edited cells, using inducible guide RNA constructs and solid wall devices to overcome growth biases and enhance editing efficiency.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If constitutively-expressed nuclease components are used to drive high efficiency editing in pooled or multiplex formats, then editing efficiency is improved, but edited cell types are rapidly depleted and unedited cells are selectively enriched
Solution Approach 1:
The patent applies preliminary action by isolating cells into single-cell suspensions or small groups before initiating genome editing. This pre-separation ensures that edited and unedited cells do not compete for survival, preventing the selective enrichment of unedited cells that occurs in bulk cultures. The isolation step is performed prior to editing, allowing each cell or small group to undergo editing independently, thereby maintaining edited cell population stability while still achieving high editing efficiency.
2Reliability
If cells are isolated into single-cell suspensions or small groups before editing, then edited cell population stability is improved, but device complexity increases due to need for isolation instruments and modules
Solution Approach 1:
The patent applies segmentation by dividing the cell population into discrete isolation units (single cells or small groups) using microfabricated devices with arrays of microwells or isolation chambers. Each unit is physically separated and can be independently processed for genome editing. This segmentation approach maintains edited cell stability while using standardized, scalable microdevice architectures that reduce overall system complexity through modular design.
Solution Approach 2:
The patent uses copying by creating multiple identical copies of the isolation device structure (arrays of microwells or chambers) that can simultaneously process many cells in parallel. This approach maintains the simplicity of a single isolation unit while achieving high throughput through replication, thereby improving edited cell stability without proportionally increasing device complexity.
3Productivity
If automated high-throughput screening and substantial isolation are implemented, then identification and enrichment of edited cells is improved, but manufacturing precision requirements increase
Solution Approach 1:
The patent applies self-service by designing isolation devices where cells automatically settle into or are trapped within microwells or isolation chambers based on their size and shape, without requiring precise manual positioning. The device structure itself provides the isolation function through geometric constraints, reducing the need for high-precision manufacturing while still achieving effective cell separation for high-throughput edited cell identification.
Data Source
AI summary
The present disclosure provides instruments, modules and methods for improved detection of edited cells following nucleic acid-guided nuclease genome editing. The disclosure provides improved automated instruments that perform methods—including high throughput methods—for screening cells that have been subjected to editing and identifying cells that have been properly edited.