Middle East Research Journal Microbiology and Biotechnology | Volume: 5 | Issue 3 | Pages: 34-49

M. R. Washikur, Md. Abdullah Al Masud, Hussain Abbas Uddin, Madia Manjum Mohona, Shomaya Akhter, Mohammad Nazir Hossain

Effective genetic diversity assessment is pivotal for sustainable fisheries management, yet standardized DNA extraction methods for non-model marine species like Harpadon nehereus (Bombay duck) remain limited. This study optimizes DNA extraction protocols to enhance the reliability of Random Amplified Polymorphic DNA (RAPD)-PCR analysis for H. nehereus from the Bay of Bengal, a vital fishery resource. Four protocols—phenol-chloroform, liquid nitrogen-assisted lysis, and two modified methods—were evaluated for DNA yield, purity, and RAPD-PCR suitability using muscle, gill, fin, and ocular tissues. DNA quality was assessed via NanoDrop spectrophotometry (A260/A280: 1.43–1.95; A260/A230: 0.49–1.93) and agarose gel electrophoresis. Fourth optimized protocol (liquid nitrogen lysis with RNase and β-mercaptoethanol) yielded the highest DNA concentration (2546.68 ± 546.60 ng/μL) with optimal purity, producing consistent RAPD-PCR banding patterns using primer OPA-03 (5′-AGTCAGCCAC-3′). Modifications, including extended incubation, Proteinase K and RNase treatment, minimized contaminants, enabling clear detection of tissue-specific and population-level genetic variations. The optimized protocol reduced technical variability, offering a reproducible, cost-effective approach for genetic fingerprinting in resource-limited settings. This work establishes a robust framework for assessing genetic diversity, detecting population bottlenecks, and informing conservation strategies for H. nehereus, contributing to sustainable fisheries management in the Bay of Bengal. These findings advance molecular ecology by providing scalable solutions for genetic studies of commercially important marine species.