Sequencing Oligonucleotide Primer for Low-Noise Index Reads

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Solution Overview

Problem

Existing sequencing methods face challenges in accurately and efficiently determining the sequence of second index sequences due to noise interference from primers attached to the solid surface and inefficiencies in sequencing order when the second read is not needed.

Innovation Solution

Employing a sequencing oligonucleotide that hybridizes to the free 3′ end of the template polynucleotide with greater affinity than the surface oligonucleotide, allowing for the sequence of the second index to be obtained prior to the paired end turn, thereby reducing noise and improving sequencing efficiency.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Measurement precision

If a sequencing primer hybridizes to a universal sequencing primer sequence on the template strand, then sequencing can be performed, but noise interference from primers attached to the solid surface reduces measurement precision

Engineering Contradiction:

Improvesequencing signal intensityVSAvoidnoise interference from surface primers

Core Design Contradiction:

Measurement precisionVSObject-affected harmful factors

Solution Approach 1:

The patent extracts the sequencing primer from the solid surface environment by using a free-floating sequencing primer that hybridizes to the template strand in solution rather than being attached to the surface. This removes the source of noise interference while maintaining the sequencing function.

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent introduces an intermediary mechanism where the sequencing primer is delivered through the liquid phase and binds to the template strand, acting as a mediator between the detection system and the template without being constrained by surface attachment. This intermediary approach eliminates direct surface-primer interactions that cause noise.

Inventive Principle:

Principle #24Intermediary (Mediator)

2Productivity

If the second index sequence is sequenced after the paired end turn using the surface oligonucleotide, then sequencing can be completed, but sequencing efficiency is reduced when the second read is not needed

Engineering Contradiction:

Improvesequencing efficiencyVSAvoidtime for unnecessary paired end turn

Core Design Contradiction:

ProductivityVSLoss of time

Solution Approach 1:

The patent performs the sequencing of the second index sequence before the paired end turn by using a free sequencing primer that can access the template strand immediately. This preliminary action allows the workflow to skip the paired end turn when the second read is not required, saving time and improving efficiency.

Inventive Principle:

Principle #10Preliminary action

Solution Approach 2:

The patent introduces dynamic flexibility to the sequencing workflow by allowing the second index sequencing to occur at different stages depending on experimental needs. The free sequencing primer enables the process to adapt and proceed without the rigid paired end turn structure when unnecessary.

Inventive Principle:

Principle #15Dynamics

3Measurement precision

If a free primer with sequence identical to surface primer is used to sequence the second index, then sequencing can be performed, but the signal intensity is insufficient due to competition with surface primer

Engineering Contradiction:

Improvesignal intensity for second index sequencingVSAvoidavailable primer for hybridization

Core Design Contradiction:

Measurement precisionVSQuantity of substance

Solution Approach 1:

The patent changes the parameters of the sequencing primer by modifying its sequence to be distinct from the surface primer sequence. This parameter change eliminates competitive hybridization while maintaining sufficient binding affinity to the template strand, thereby improving signal intensity.

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The patent applies local quality differentiation by designing the sequencing primer with a specific sequence modification at the region that interacts with the template strand. This localized sequence differentiation ensures specific binding without competition from surface primers, enhancing signal quality.

Inventive Principle:

Principle #3Local quality

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 sufficiently intense signal for sequencing the second index sequence without noise interference, enabling more efficient sequencing when the second read is not required.

Implementation Method 1

a sequencing oligonucleotide that hybridizes to the free 3′ end of the template polynucleotide with greater affinity than the surface oligonucleotide

Methodology Applied

Scientific EffectHybridization:

Implementation Method 2

extending the sequencing oligonucleotide using the template polynucleotide as a template

Methodology Applied

Scientific EffectDNA synthesis:

Data Source

PatentUS12529098B2Primer oligonucleotide for sequencing

Publication Date: 2026.01.20 ILLUMINA CAMBRIDGE LTD

AI summary

Polynucleotide sequencing methods employ a sequencing oligonucleotide that hybridizes to a free 3′ end potion of a template polynucleotide strand with greater affinity than a surface oligonucleotide. Such sequencing oligonucleotides may be used as a primer to determine the sequence of an index sequence by extending the sequencing oligonucleotide using the template strand as a template. Sequencing processes that employ such sequencing oligonucleotides provide a sufficiently intense signal to determine to the sequence of the index sequence.