GoFish: A versatile nested PCR strategy for environmental DNA assays for marine vertebrates.

Area of Science:

• Environmental DNA (eDNA) analysis
• Molecular ecology
• Biodiversity monitoring

Background:

• Environmental DNA (eDNA) is a powerful tool for species detection.
• Existing methods like metabarcoding and qPCR have limitations in cost, speed, and accessibility.
• There is a need for accessible, rapid, single-species eDNA assays.

Purpose of the Study:

• To develop and validate GoFish, a novel strategy for single-species eDNA presence/absence assays.
• To assess the performance of GoFish compared to existing eDNA methods.
• To demonstrate the utility of GoFish for monitoring aquatic species.

Main Methods:

• Utilized nested PCR with species-specific primers targeting mitochondrial 12S rDNA.
• Developed assays for 11 bony fish, 9 cartilaginous fish, and bottlenose dolphin.
• Validated sensitivity against Illumina MiSeq metabarcoding and assessed practical workflow.

Main Results:

• GoFish demonstrated sensitivity comparable to Illumina MiSeq metabarcoding.
• The assay successfully detected target species in environmental samples.
• GoFish offers a faster (3-day) and less technically demanding alternative to other methods.

Conclusions:

• GoFish is a sensitive, rapid, and accessible method for single-species eDNA presence/absence detection.
• It overcomes limitations of qPCR and Illumina metabarcoding, requiring only basic equipment.
• GoFish is suitable for species monitoring, educational purposes, and situations requiring quick results.