Molecular Identification of Residual DNA Separated from the Persian Sturgeon (Acipenser persicus) for Modeling eDNA Evaluation in Aquatic Ecosystem

Abstract

Considering the importance of Caspian Sea sturgeon conservation according to CITES rules, finding effective and efficient methods for tracing and identifying sturgeon species are necessary. Residual DNA detection in fish ponds can be used as a model for tracing fish environmental DNA (eDNA) in rivers and seas. This method of DNA detection is non-invasive and advantageous for the conservation of critically endangered sturgeons. Sampling was done from the water of the Persian sturgeon fish pond and fixed with precipitation premix solution (ethanol and acetate sodium). DNA was extracted from fixed water, and fine tissue of two sturgeon species in the Caspian Sea, Sterlet, and Siberian sturgeon. The positive specificity of primers was checked by conventional PCR for sturgeons with fine tissue DNA samples. The linear relationship between the threshold cycle (ct) value and the Persian Sturgeon DNA concentration was measured by the Mini-Barcoding quantitative real-time PCR. DNA samples of fish ponds generated a curve that could not be produced in negative control and irrelevant (non-sturgeon) genomic DNA. Although residual DNA of Persian sturgeons was detected in fish pond water at picogram level through the quantitative method, molecular diagnosis with this method can confirm only the existence of sturgeon species (in general) in fish ponds. In identifying residual DNA in the picogram level of the Persian sturgeon fish pond, SYBER Green real-time PCR method can be an acceptable method for barcoding with high efficiency compared to other methods such as conventional PCR method and non-invasive which is advantageous for the conservation of critically endangered sturgeons.

Keywords

• aquatic ecosystem
• environmental dna
• persian sturgeon
• real-time pcr