Nonrandom Oligonucleotide Adapters for Sequencing Error Reduction
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Solution Overview
Problem
Current methods for determining nucleotide sequences in nucleic acid samples are prone to sequencing errors and lack sensitivity in detecting low-frequency genetic alterations, particularly in identifying genetic variations that can lead to medical conditions.
Innovation Solution
The use of nonrandom oligonucleotide adapters with predetermined molecular barcode sequences is introduced, which are ligated to nucleic acid templates, allowing for efficient sequencing and amplification, thereby reducing errors and increasing sensitivity in detecting genetic alterations by providing a streamlined approach to automated sequencing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional sequencing methods are used, then sequencing can be performed, but sequencing errors occur and sensitivity for detecting low-frequency genetic alterations is insufficient
Solution Approach 1:
The adapter is segmented into distinct functional regions: a first region that anneals to the template and a second region containing the molecular barcode. This segmentation allows the barcode to be separately optimized for error correction while the annealing region maintains template binding, thereby improving sequencing accuracy without compromising detection sensitivity
Solution Approach 2:
The molecular barcode in the adapter acts as an intermediary that mediates error correction. The barcode sequences are designed to correct sequencing errors and enable detection of low-frequency genetic alterations by serving as a reference against which template sequences can be compared, thus resolving the contradiction between reliability and measurement precision
2Reliability
If nonrandom oligonucleotide adapters with molecular barcodes are used, then sequencing errors are reduced and detection sensitivity is improved, but adapter design and manufacture complexity increases
Solution Approach 1:
The adapter design incorporates universal functional elements that can be reused across different sequencing applications. The first region that anneals to the template and the second region with the molecular barcode are designed to work together in a standardized format, reducing design complexity while maintaining improved sequencing accuracy and detection sensitivity
Solution Approach 2:
The adapter design uses specific parameter optimizations: the molecular barcode is configured with 1-5 nucleotides in the second region, and the annealing region in the first region is optimized for template binding. These parameter changes establish a balanced design that achieves high reliability without excessive complexity
3Measurement precision
If molecular barcodes are incorporated into adapters, then sequencing error reduction and detection sensitivity improvement are achieved, but manufacturing process complexity increases
Solution Approach 1:
The molecular barcodes are pre-incorporated into the adapter sequences during the adapter synthesis stage, rather than being added separately during sample processing. This preliminary action simplifies the manufacturing process by integrating the error-correction functionality directly into the adapter oligonucleotides, making them ready-to-use while maintaining high detection sensitivity
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
This method significantly reduces sequencing errors and enhances the detection of low-frequency single nucleotide alterations, providing a more accurate and efficient means to determine nucleotide sequences and diagnose genetic conditions.
Implementation Method 1
the polynucleotide B species are annealed to complementary polynucleotide B′ species
Data Source
AI summary
Technology provided herein relates in part to methods, processes, machines and apparatuses for determining sequences of nucleotides for nucleic acid templates in a nucleic acid sample. The technology provide herein also relates in part to methods, processes, machines and apparatuses for counting nucleic acid templates. Nucleic acid templates of a sample are tagged with nonrandom oligonucleotide adapters that include predetermined non-randomly generated sequences. The use of these nonrandom oligonucleotide adapters provides an efficient method to reduce sequencing errors, and increase the sensitivity of detection of low-frequency single nucleotide alterations.