DNA Spike-Ins for Contamination Tracking in Amplicon Sequencing
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Solution Overview
Problem
Amplicon-based genome sequencing methods face high risks of contamination during amplification, particularly in large-scale viral genomic surveillance, which can confound genomic analyses and undermine the integrity of SARS-CoV-2 genomes, especially in low-titer samples and when performed by laboratories with limited experience.
Innovation Solution
The implementation of synthetic DNA spike-ins (SDSIs) with unique sequences in each sample, capable of amplification and differentiation, to detect and prevent contamination by identifying sample swaps and cross-contamination through sequencing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If amplicon-based sequencing methods are used to accelerate large-scale viral genomic surveillance, then sequencing speed and sensitivity are improved, but the risk of contamination during amplification increases significantly
Solution Approach 1:
The patent introduces synthetic DNA spike-ins as intermediary control elements that are added to each sample before amplification. These spike-ins serve as mediators to monitor and detect contamination events during the high-speed amplicon-based sequencing process, allowing laboratories to maintain high productivity while identifying contamination risks through the presence of unexpected spike-in sequences in sequencing results
Solution Approach 2:
The patent implements a feedback mechanism where the amplification and sequencing of synthetic DNA spike-ins provides real-time information about contamination events. By analyzing the sequencing results for spike-in sequences that should not be present, the system generates feedback that indicates whether contamination has occurred, allowing for immediate identification and correction of contamination issues while maintaining high sequencing throughput
2Productivity
If multiple large batches of samples are processed in parallel to meet surveillance goals, then productivity increases, but the potential for contamination increases
Solution Approach 1:
The patent divides the surveillance workflow into segmented batches, each with its own unique synthetic DNA spike-in control set. By segmenting the control elements along with the samples and processing them in parallel batches, the system can maintain high throughput while ensuring that contamination in one batch does not compromise other batches, as each batch has its own dedicated spike-in identifiers for tracking
Solution Approach 2:
Unique synthetic DNA spike-in sequences serve as intermediary tracking elements for each batch and sample. These mediators allow parallel processing of multiple batches while maintaining the ability to identify and trace contamination sources, ensuring data integrity across high-throughput surveillance operations by providing batch-specific control signals in the sequencing output
3Measurement precision
If synthetic DNA spike-ins with unique sequences are added to each sample for contamination detection, then measurement precision is improved, but device complexity increases
Solution Approach 1:
The patent merges the contamination detection function with the existing amplification and sequencing workflow by using the same PCR primers and sequencing infrastructure. The synthetic DNA spike-ins are designed to be amplified alongside target viral sequences using the same reagents and equipment, combining multiple functions (contamination monitoring, sample tracking, and viral detection) into a unified protocol that leverages existing laboratory capabilities without requiring separate complex systems
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
SDSIs enhance the reliability of genomic data by detecting and correcting errors, ensuring accurate sample identification and improved genome recovery without impacting sequencing efficiency, suitable for diverse laboratories and clinical applications.
Implementation Method 1
amplifying one or more of the cDNA samples and SDSI
Data Source
AI summary
Embodiments disclosed herein provide methods of using synthetic DNA spike-ins (SDSIs) to detect, prevent, and quantify contamination in amplicon sequencing. These embodiments may, but are not limited to, reveal sample swaps, intra-batch contamination, and, on a larger scale, intra-laboratory contamination. Embodiments disclosed herein also provide synthetic DNA spike-ins for use in amplicon-based sequencing methods.