Nuclease-Resistant Primer Sets for Reusable Sequencing Flow Cells
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Solution Overview
Problem
Existing nucleic acid detection methods often require single-use flow cells with surface chemistry, limiting their reusability and increasing costs.
Innovation Solution
Development of reusable primer sets and flow cells that allow multiple uses with different samples, utilizing nuclease-resistant anchor oligonucleotides and regenerable primer sets to maintain functionality.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional surface chemistry is used in flow cells, then detection function is achieved, but the flow cell can only be used once and must be discarded
Solution Approach 1:
The primer set is divided into two functional parts: a stable anchor oligonucleotide that remains immobilized on the flow cell surface, and a consumable primer portion that is depleted during sequencing. This segmentation allows the anchor to be reused while the consumable portion is replaced, resolving the contradiction between maintaining detection function and enabling reusability.
Solution Approach 2:
The patent implements a system where the consumable primer portion is discarded after use (depleted during sequencing), while the anchor oligonucleotide is recovered and retained on the flow cell surface for subsequent uses. This selective discarding and recovering enables the flow cell to be reused multiple times while maintaining detection functionality.
2Measurement precision
If single-use flow cells are used, then detection accuracy is maintained, but waste and costs increase
Solution Approach 1:
By segmenting the primer set into a reusable anchor and a consumable portion, the patent allows the valuable anchor component to be preserved and reused, thereby reducing waste while maintaining detection accuracy through the consistent presence of the anchor on the flow cell surface.
Solution Approach 2:
The consumable primer portion is discarded after depletion, while the anchor oligonucleotide is recovered and retained on the flow cell. This selective recovery of the anchor reduces material waste and costs while maintaining the detection precision provided by the stable anchor presence.
3Productivity
If primers are immobilized on flow cell surface, then sequencing reaction occurs, but primers are depleted and flow cell cannot be reused
Solution Approach 1:
The primer set is segmented into a permanent anchor oligonucleotide that remains immobilized on the flow cell surface and a consumable primer portion that participates in the sequencing reaction. This segmentation allows the sequencing reaction to proceed using the consumable portion while the anchor remains available for subsequent reactions, extending the flow cell usage duration.
Solution Approach 2:
After the sequencing reaction depletes the consumable primer portion, this portion is discarded while the anchor oligonucleotide is recovered and retained on the flow cell surface. This recovery of the anchor enables the flow cell to be reused for additional sequencing reactions, extending its operational duration.
Data Source
Figure 1A~1C
Figure 2~3E
Figure 4~5D
AI summary
An example primer set includes first and second nuclease resistant primers. The first nuclease resistant primer includes a first sequence; a first cleavage site attached at a 3' end of the first sequence; and a first nuclease resistant modification incorporated between the first sequence and the first cleavage site. The second nuclease resistant primer includes a second sequence that is different from the first sequence; a second nuclease resistant modification incorporated at a 3' end of the second sequence; and a second cleavage site attached between the second sequence and the second nuclease resistant modification. The second cleavage site is different from the first cleavage site.