DNA Origami Nanostructures for Paired TCR Sequencing
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current methods are unable to obtain linked TCR alpha and beta CDR3 sequence information from individual cells within large mixed populations, as existing technologies either require costly single cell sorting or result in scrambled genomic DNA and mRNA, precluding paired analysis.
Innovation Solution
The development of DNA origami nanostructures that bind, isolate, and link TCR alpha and beta CDR3 mRNA within individual cells, allowing for high-efficiency transfection, isolation, and deep sequencing of paired CDR3 sequences using DNA origami scaffolds and complementary staple sequences.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If single cell sorting is used to obtain TCR alpha and beta CDR3 sequences from individual cells, then linked CDR3 information can be obtained, but the cost and complexity increase significantly
Solution Approach 1:
The patent segments the TCR alpha and beta mRNA molecules by attaching them to separate DNA origami nanostructures, each carrying specific molecular barcodes. This segmentation allows individual cell analysis without physical sorting, as each segmented unit retains unique identification markers for later reconstruction of paired CDR3 information.
Solution Approach 2:
DNA origami nanostructures serve as intermediary carriers that bind to TCR alpha and beta mRNA molecules. These nanostructures contain molecular barcodes that act as intermediaries to track and link the pairing information between alpha and beta chains from individual cells, eliminating the need for single cell sorting while preserving linked CDR3 information.
2Ease of manufacture
If mixed cell populations are lysed to obtain CDR3 sequences, then processing is simplified, but genomic DNA and mRNA become scrambled precluding paired analysis
Solution Approach 1:
The patent performs preliminary action by attaching molecular barcodes to TCR alpha and beta mRNA molecules before cell lysis. The DNA origami nanostructures are introduced and bound to the mRNA in intact cells, establishing the linkage information in advance. This preliminary tagging allows subsequent bulk lysis processing while preserving the paired CDR3 information through the pre-established barcode connections.
Solution Approach 2:
The patent creates informational copies through molecular barcodes on DNA origami nanostructures that replicate the pairing information of TCR alpha and beta chains. Instead of relying on the physical integrity of mRNA molecules through lysis, the barcode copies preserve the linkage information, allowing simplified bulk processing while maintaining the ability to reconstruct paired CDR3 sequences.
3Productivity
If current methods are used to analyze TCR repertoires, then analysis is possible, but linked CDR3 information from individual cells cannot be obtained
Solution Approach 1:
The patent adds another dimension to TCR repertoire analysis by introducing DNA origami nanostructures with molecular barcodes that provide individual cell resolution within bulk populations. This dimensional addition allows simultaneous high-throughput processing of many cells while maintaining the ability to resolve and analyze linked CDR3 information at the individual cell level, overcoming the traditional trade-off between productivity and 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 the efficient analysis of TCR repertoires without the need for single cell sorting, providing linked CDR3 sequence information for large populations, thereby enhancing the understanding of immune diversity and competency.
Implementation Method 1
The specificity of the interactions between complementary base pairs make DNA a useful construction material
Data Source
AI summary
Methods for construction of DNA origami nanostructures, as well as for binding, isolation, linking, and deep sequencing information, such as both of TCR alpha and beta CDR3 mRNA, from individual cells within a mixed population of cells without the need for single cell sorting (FIG. 1).