Background

Drosophila melanogaster, a holometabolous, invertebrate model organism, has contributed extensively towards analyzing the roles of various phylogenetically conserved genes (Green et al., 2018; Dubey et al., 2020). Toxicological studies utilizing flies have also aided in the assessment of the effects of heavy metals that interfere with important biological processes (Nanda and Firdaus, 2021; Nanda and Firdaus, 2022), including immune system (Nanda et al., 2019). Drosophila research has also paved a way for the exploration of innumerable scientific questions in various branches of biological sciences. Notably, Drosophila melanogaster genome is 60% homologous to humans (Adams et al., 2000).

Analysis of Drosophila health and activity is a fundamental aspect to quantify any adversities either due to genetic manipulation or dietary exposure to toxicant. Among the various life traits, quantifying stagewise lethality, checking life span (Carey et al., 2006), and examining locomotor and reproductive capabilities (Rogina et al., 2007) are the biomarkers most often used (Landis et al., 2020; Olcott et al., 2010). Several behavioral assays also facilitate the analysis of a typical mutant from embryonic to adult stage (Ain and Firdaus, 2021).

The Cu, Zn superoxide dismutase (SOD1) is an antioxidant enzyme that provides protection against excessive reactive oxygen species (ROS) in Drosophila melanogaster. The three isoforms of SOD in Drosophila, namely SOD1, mitochondrial Mn superoxide dismutase SOD 2 (SOD2), and extracellular Cu Zn superoxide dismutase (SOD 3), are encoded by genes located on separate chromosomal loci (Duttaroy et al., 1994; Blackney et al., 2014). In this protocol, we used Drosophila wherein the cytosolic SOD1 gene expression was either overexpressed or downregulated in a ubiquitous manner through genetic crosses between the driver Act-5c-GAL4 and the responder RNAi/overexpression fly lines. This enabled the study of the role of SOD1 gene in life span, lethality, and motility behaviour of Drosophila.

We describe a simple continuous method that requires a single experimental setup to perform both the lethality and life span assays of the genetically altered or toxicant-exposed Drosophila. The protocol is unique and significant, due to the fact that it highlights the usage of minimal laboratory equipment and uses one single experimental setup for continuous monitoring of lethality followed by survival analysis and crawling and climbing assays in larvae and adults, respectively.