Library Preparation Adapters With UMIs for Low-Input Sequencing
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Solution Overview
Problem
Existing sequencing technologies face challenges in efficiently processing biological samples with high throughput and maximizing the use of limited genetic material, such as DNA and RNA, for library preparation.
Innovation Solution
The development of non-naturally occurring nucleic acid molecules with specific sequences (e.g., SEQ ID Nos: 1-1264) and adapters with strategic mismatches, coupled to supports like beads, for efficient library preparation, including methods for generating unique barcode sequences and hybridization-based coupling.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional library preparation methods are used, then the process can be completed with standard protocols, but the efficiency is low and large amounts of genetic material are required
Solution Approach 1:
The patent modifies the chemical structure of adapters by introducing strategic mismatches at specific positions (e.g., position 10-15 in the P5 region and position 10-15 in the P7 region) compared to conventional adapters. These parameter changes in adapter sequence composition enable more efficient binding to template nucleic acids, thereby improving library preparation efficiency while reducing the quantity of genetic material required.
Solution Approach 2:
The patent employs specially designed adapters as intermediary molecules that facilitate the binding between template nucleic acids and sequencing platforms. These adapters include unique molecular identifiers (UMIs) and barcode sequences that act as mediators to enable efficient library preparation and sample identification, resolving the contradiction between productivity and material quantity requirements.
2Productivity
If high throughput sequencing is implemented, then more samples can be processed, but the complexity of sample identification and tracking increases
Solution Approach 1:
The patent segments sample identification into multiple components: unique molecular identifiers (UMIs), barcode sequences, and index sequences. Each component serves a specific function in tracking and identifying samples during high-throughput sequencing, thereby managing complexity while maintaining high productivity.
Solution Approach 2:
The adapters designed in the patent serve multiple functions simultaneously: they bind to template nucleic acids, provide unique molecular identifiers for tracking, include barcode sequences for sample identification, and enable efficient library preparation. This multi-functionality reduces the overall complexity of sample identification in high-throughput sequencing.
3Quantity of substance
If limited genetic material is available, then sample scarcity is addressed, but the ability to perform comprehensive analysis is reduced
Solution Approach 1:
The patent employs unique molecular identifiers (UMIs) that create molecular copies with distinct identification sequences. These copies allow comprehensive analysis of limited genetic material through multiple sequencing reads, enabling complete characterization of rare or limited samples while using minimal genetic material.
Solution Approach 2:
The adapters are pre-designed with strategic mismatches and unique molecular identifiers before the sequencing process. This preliminary action enables efficient binding and identification of limited genetic material, allowing comprehensive analysis to be performed on small sample quantities without requiring large amounts of genetic material.
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
Enhances the efficiency of library preparation by maximizing the use of limited genetic material and enabling high-throughput sequencing, while ensuring unique identification of samples.
Implementation Method 1
the coupling is via ligation
Implementation Method 2
the coupling comprises hybridization
Data Source
AI summary
Provided herein are systems, methods, compositions, and kits for library preparation. In some cases, multiple distinct types of adapter molecules may be provided to a template nucleic acid molecule. In some cases, a single type of adapter molecule may be provided to a template molecule. In some cases, multiple distinct types of adapter molecules may be sequentially provided to a template molecule to form multi-adapter template complexes.