Spatially Barcoded Microarray with Length-Encoded Probe Locations
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Solution Overview
Problem
Current spatial barcoding technologies for microarrays in molecular biology, such as microspotting, microfluidic channels, and split-pool-barcoded beads, suffer from limitations like low resolution, high cost, complexity, and the need for time-consuming decoding processes.
Innovation Solution
A spatially barcoded microarray is developed using oligonucleotides with varying lengths to determine probe locations, allowing for easy fabrication, flexible array dimensions, and scalable resolution, achieved through controlled enzymatic removal of nucleotides in a fluidic flow.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If microspotting or microfluidic channels are used for spatial barcoding, then the microarray can be fabricated, but the resolution is limited
Solution Approach 1:
The patent changes the parameter of oligonucleotide length to encode spatial information. By varying the length of barcode oligonucleotides (e.g., 20-40 nucleotides) instead of using fixed-length sequences, the system achieves high spatial resolution through length-based differentiation while maintaining simple fabrication processes.
Solution Approach 2:
The patent replaces mechanical positioning systems (microspotting, microfluidic channels) with a biochemical approach using enzymatic processing. By using exonuclease treatment duration to control oligonucleotide length, the system substitutes mechanical precision requirements with temporal control of a chemical process, achieving high resolution without complex fabrication machinery.
2Measurement precision
If split-pool-barcoded beads are used, then higher resolution is achieved, but the decoding process becomes time-consuming and requires costly equipment
Solution Approach 1:
The patent extracts the decoding complexity from the experimental workflow by embedding spatial information directly into the oligonucleotide length during fabrication. This eliminates the need for separate decoding processes and expensive sequencing equipment, as the spatial barcode is inherently encoded in the physical dimension of the oligonucleotide.
Solution Approach 2:
The patent uses simple, inexpensive oligonucleotide probes with varying lengths as disposable barcodes. These short-lived molecular objects carry spatial information through their length, eliminating the need for expensive, complex decoding equipment and reducing experimental time requirements compared to persistent bead-based systems.
3Reliability
If in-situ solid-phase amplification is used, then spatial barcoding is achieved, but the process becomes complicated and expensive
Solution Approach 1:
The patent segments the probe structure into distinct functional regions: a location-specific barcode oligonucleotide region with varying length, a capture region, and a unique identifier. This segmentation allows each region to perform its specific function independently, simplifying the overall process while maintaining reliability through modular design.
Solution Approach 2:
The patent performs the spatial barcoding action during the fabrication process itself by controlling exonuclease treatment duration to create probes of specific lengths. This preliminary action encodes spatial information before the probes are used in experiments, eliminating the need for complex in-situ amplification procedures and reducing overall process complexity.
4Adaptability or versatility
If conventional spatial barcoding methods are used, then microarrays can be fabricated, but they lack flexibility in array dimension and resolution
Solution Approach 1:
The patent introduces dynamic adjustability in array dimension and resolution through the length of barcode oligonucleotides. By varying the length parameter (e.g., 20-40 nucleotides) during fabrication, the system can adapt to different array dimensions and resolution requirements without changing the fundamental fabrication process, achieving versatility while maintaining ease of manufacture.
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
The method provides a cost-effective, easily fabricated, and highly scalable microarray with improved resolution and flexibility, enabling precise determination of nucleic acid targets in spatial transcriptomics.
Implementation Method 1
exposing the plurality of the first oligonucleotides to a fluidic flow, wherein the fluidic flow comprises a first liquid and a second liquid that flow along a first direction, wherein the first liquid and the second liquid are immiscible with each other and form an interface parallel to a first direction, wherein the first liquid comprises a first enzyme capable of removing one or more nucleotides from the first oligonucleotides
Implementation Method 2
the first liquid and the second liquid are immiscible with each other and form an interface parallel to a first direction
Implementation Method 3
exposing the plurality of the first oligonucleotides to a fluidic flow, wherein the fluidic flow comprises a first liquid and a second liquid that flow along a first direction
Data Source
AI summary
The present disclosure provides a microarray comprising a plurality of probes. Each probe comprises a first oligonucleotide and a second oligonucleotide. The location of each probe on the microarray can be determined by the length of the first oligonucleotide and the length of the second oligonucleotide, thus providing a spatially barcoded microarray. Also provided are the methods of producing such spatially barcoded microarray. Also provided are the method of using such spatially barcoded microarray.