Double-End Library Tags for MGI Sequencing Crosstalk Reduction
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Solution Overview
Problem
The MGI sequencing platform faces challenges with sample crosstalk due to the use of single-end library tags, leading to data contamination and reduced sequencing data splitting rates, which can be exacerbated by issues in adapter synthesis and sequencing processes.
Innovation Solution
The implementation of double-end library tags with standardized lengths and base compositions, specifically 5' and 3' end tags with balanced GC content and minimal continuous same bases, to enhance base balance and reading accuracy, thereby improving sequencing data splitting.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If double-end library tags are used to reduce sample crosstalk, then sample crosstalk is reduced, but sequencing data splitting rate decreases due to reading accuracy problems
Solution Approach 1:
The patent changes the parameters of library tags from single-end to double-end configuration, with specific constraints on tag length (8-12 bp), GC content (40-60%), and base composition balance. These parameter optimizations ensure that while using double-end tags to reduce crosstalk, the sequencing machine can still accurately read the tags and maintain high data splitting rates.
Solution Approach 2:
The patent creates a composite library tag system combining 5' end tags and 3' end tags with complementary base compositions. The 5' tag and 3' tag are designed to work together as a pair, where their combined base compositions are balanced, improving both crosstalk reduction and reading accuracy simultaneously.
2Reliability
If double-end library tags are used to eliminate crosstalk problems, then crosstalk is eliminated, but sequencing cost increases due to reduced splitting rate
Solution Approach 1:
By optimizing tag length to 8-12 bp and GC content to 40-60%, the patent ensures that double-end tags can be read accurately by the sequencing machine, maintaining high data splitting rates and avoiding the cost increase that would result from low splitting rates.
Solution Approach 2:
The patent replaces the single-end tag mechanism with a double-end tag mechanism that uses complementary base composition design. This substitution enables the system to achieve both crosstalk elimination and cost efficiency by maintaining high data splitting rates through improved tag readability.
3Device complexity
If single-end library tags are used for sequencing, then sequencing process is simple, but sample crosstalk occurs due to adapter or primer contamination
Solution Approach 1:
The patent segments the single library tag into two separate tags: a 5' end tag and a 3' end tag. Each tag is independently designed with balanced base compositions, and both are required for successful data splitting. This segmentation eliminates crosstalk because contamination affecting only one tag will not cause misassignment, as both tags must match for correct sample identification.
Data Source
AI summary
The invention provides a double-end library tags composition and application thereof in MGI sequencing platform. The double-end library tags composition includes a plurality of 5′-end library tags and a plurality of 3′-end library tags, the lengths of the plurality of 5′-end library tags are all the same, the lengths of the plurality of 3′-end library tags are all the same, and in the double-end library tags composition, the occurrences of each base at the same position are also all the same.