Rapid DNA release from arthropod specimens using the STEP buffer for molecular diagnostics

Abstract

Rapid and reliable release of genomic DNA (gDNA) from arthropod specimens is essential for molecular diagnostics, species identification, and ecological monitoring, particularly when only minute samples or limited body parts are available. Here, we present the STEP buffer, a newly formulated DNA release solution, and compare its performance with that of the previously reported DAPE buffer and water, using whole bodies of Dermatophagoides pteronyssinus and Cimex lectularius first-instar nymph, exuviae of C. lectularius, and hind legs of Aedes albopictus. Specimens were incubated in each buffer at 95°C for 5 min. The supernatant was subsequently used directly as the gDNA template for quantitative PCR, as well as for downstream compatibility testing by PCR and two types of loop-mediated isothermal amplification (LAMP) assays. The STEP buffer yielded gDNA amounts comparable to or greater than those obtained using DAPE. In addition, it facilitated complete submersion of hydrophobic arthropod cuticles during incubation. PCR assays confirmed high amplification success rates for STEP, particularly from specimens containing limited gDNA. In conventional LAMP, all buffer types supported amplification. In contrast, in colorimetric LAMP, only STEP maintained consistent detection, while DAPE caused false negatives due to pH interference. Thus, the STEP buffer provides a rapid, robust, and broadly compatible method for DNA release, enabling reliable PCR and LAMP detection even from minute arthropod samples, which is advantageous for on-site diagnostics and applications requiring specimen preservation.