CRISPR-Cas Cell Sorting via DNA Barcode Targeting
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current methods for isolating specific cell populations from genetically heterogeneous cell populations are laborious, slow, and of low throughput, and do not allow for determining gene and protein expression before drug treatment or recovering pure populations of interest.
Innovation Solution
A method using CRISPR-Cas systems with DNA barcodes and selection markers to sort cells based on specific genetic targets, allowing for the recovery of cells with specific DNA barcodes from a heterogeneous population by activating or inactivating selection markers through targeted insertion or deletion.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If traditional methods for isolating specific cell populations are used, then cells can be separated, but the process is laborious, slow, and of low throughput
Solution Approach 1:
The patent replaces traditional mechanical/manual cell sorting methods with an automated optical sorting system. Cells are identified and sorted based on their fluorescent signal characteristics using flow cytometry or similar optical sorting technologies, eliminating the need for manual intervention and significantly increasing throughput while reducing labor requirements
Solution Approach 2:
The patent utilizes fluorescent markers that cause cells to emit light at specific wavelengths. Cells expressing the fluorescent marker are detected and sorted based on their fluorescent signal intensity and characteristics, enabling automated identification and separation of specific cell populations from heterogeneous mixtures
2Loss of information
If traditional cell isolation methods are used, then cells can be separated, but gene and protein expression cannot be determined before drug treatment
Solution Approach 1:
The patent incorporates fluorescent markers or other detectable signals into cells before drug treatment begins. These markers allow researchers to detect and analyze gene and protein expression levels in sorted cell populations prior to treatment, preserving the ability to determine expression information without delaying the treatment protocol
Solution Approach 2:
The patent uses fluorescent markers or signal molecules as intermediaries that carry information about cell state and expression levels. These markers can be detected by flow cytometry or other optical methods, enabling non-destructive measurement of gene and protein expression before drug treatment without interfering with subsequent treatment efficacy
3Reliability
If traditional cell sorting methods are used, then cells can be isolated, but pure populations of interest cannot be recovered
Solution Approach 1:
The patent employs automated optical sorting systems that use fluorescent signal detection to identify and isolate pure cell populations. The system can distinguish cells based on their fluorescent characteristics and physically separate them with high purity, eliminating the need for manual sorting and enabling recovery of genetically homogeneous populations
Solution Approach 2:
The patent utilizes fluorescent markers that provide distinct optical signals for cell identification and sorting. Cells expressing the fluorescent marker emit light at specific wavelengths that can be detected and used to sort pure populations, enabling reliable recovery of cells with desired genetic characteristics through optical detection and separation
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 non-destructive sorting and isolation of specific cell populations with increased specificity, facilitating the recovery of cells with desired genetic characteristics for further analysis.
Implementation Method 1
Cas9 forms a complex with a small guide RNA that directs the enzyme to its DNA target via Watson-Crick base pairing
Implementation Method 2
the CRISPR-Cas complex comprising the CRISPR-Cas enzyme complexed with the CRISPR-Cas guide RNA and the DNA barcode of interest, thereby activating transcription of the selection marker or reporter
Data Source
AI summary
The invention provides for use of CRISPR-Cas systems to sort barcoded cells or molecules. Cells or nucleic acid molecules may be sorted from a heterogenous population by targeting a barcode of interest specific for a cell or cell progeny or nucleic acid molecule of interest.