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FishChip: Development of a High-Throughput qPCR Assay for Freshwater Fish eDNA Quantification and Its Comparison With
Shoutao Cheng1,2, Jiasheng Zhang1, Zhuming Liang1
1State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, Beijing, People's Republic of China.
Molecular Ecology Resources
|August 5, 2026
Summary
High-throughput quantitative PCR (HT-qPCR) offers a scalable solution for environmental DNA (eDNA) fish monitoring. This method shows strong correlation with droplet digital PCR (ddPCR), enabling efficient basin-scale aquatic ecosystem assessments.
Area of Science:
- Environmental DNA (eDNA) analysis
- Aquatic ecosystem monitoring
- Molecular biology techniques
Background:
- Quantitative monitoring of fish populations using eDNA is crucial for basin-scale surveys.
- Traditional methods like quantitative real-time PCR (qPCR) and droplet digital PCR (ddPCR) face throughput limitations.
- High-throughput qPCR (HT-qPCR) presents a potential solution for increased sample and target processing capacity.
Purpose of the Study:
- To evaluate the performance and comparability of HT-qPCR for fish eDNA quantification against ddPCR.
- To assess the utility of HT-qPCR for large-scale aquatic ecosystem monitoring.
- To provide methodological insights for eDNA-based fish assessment.
Main Methods:
- Designed 19 species-specific primer pairs for nine freshwater fish species.
- Validated primer performance on a high-throughput chip-based qPCR platform.
- Quantified fish eDNA across six aquatic regions using HT-qPCR and compared results with ddPCR.
Main Results:
- All designed assays performed well on the HT-qPCR platform, with high amplification efficiencies.
- HT-qPCR achieved a 100% detection rate for Cyprinus carpio (C. carpio).
- HT-qPCR and ddPCR estimates showed strong correlation (r=0.858) and near-equivalence, with HT-qPCR showing a ~3-fold overestimation.
Conclusions:
- HT-qPCR is a validated and effective tool for catchment-scale fish eDNA quantification.
- Methodological innovations and technical support are provided for aquatic ecosystem monitoring.
- A correction factor may harmonize data between HT-qPCR and ddPCR for consistent assessments.
