[Objectives] Sturgeons are evolutionarily significant and economically important fish in China, particularly valued for caviar production. Due to the lack of distinct secondary sexual characteristics between sexes, sex identification of farmed sturgeons remains challenging with conventional methods. This study aims to develop a minimally invasive DNA-based molecular marker for accurate early sex identification in farmed sturgeons, providing a solution that is minimally invasive, rapid, accurate, species-versatile, and beneficial for both sturgeon aquaculture and conservation efforts. [Methods] A total of 155 sturgeon samples were collected from Zhejiang and Heilongjiang provinces. The 110 samples from Zhejiang, with known sex information, served as standard controls, while the 45 samples from Heilongjiang were juvenile sturgeons of unknown sex. DNA was extracted from the dorsal fin tissue via a commercial kit. An ultra-trace nucleic acid protein analyzer was used to measure the DNA purity and concentration. Specific primers (Asc418F/Asc418R) were designed by Primer Premier 5.0 software based on the genome resequencing data from 3-year-old Acipenser schrenckii. Temperature gradient PCR (45 ~ 62 ℃) was performed to optimize the annealing temperature, and the primer sensitivity was verified via a 10-fold serial dilution of plasmid DNA. [Results] The female-specific 418-bp DNA fragment (designated ZXJAscF418) was successfully amplified only in female sturgeon samples. The results demonstrated that the primers for this DNA marker exhibited excellent amplification specificity, with an effective annealing temperature range of 45 ~ 59 ℃ (Fig. 1). The optimal annealing temperature was 56 ℃, as band intensity decreased above 58 ℃ and became undetectable at 62 ℃. The primers exhibited high sensitivity, reliably detecting DNA concentrations as low as 6.55 × 10^-6 mg/L, with complete loss of visible bands at a 10^-8 dilution (Fig. 2). Application of this molecular marker achieved 100% accuracy in sex identification for four sturgeon species (A. schrenckii, Huso dauricus, H. dauricus ♀ × A. schrenckii ♂ F1, and A. gueldenstaedtii) from Zhejiang (Fig. 3). For the juvenile sturgeons of unknown sexes from Heilongjiang, the method predicted a female-to-male ratio of 1.25︰1 (Fig. 4). [Conclusion] This study developed a DNA-based molecular marker assay for sex identification in farmed sturgeons. The assay demonstrated minimally invasive sampling, operational simplicity, high rapidity, and high accuracy across multiple sturgeon species. This approach enables reliable early sex identification, with significant implications for sturgeon health management, cost saving, and conservation efforts. It may serve as a critical tool for restoring endangered sturgeon populations through artificial breeding.