Accurate identification of butterfly species is a fundamental requirementfor biodiversity research, ecological monitoring, and conservationplanning. Traditionally, butterfly taxonomy has relied heavily onmorphological features such as wing coloration, size, and pattern.However, these characteristics are often insufficient due topolymorphism, sexual dimorphism, mimicry, and cryptic species, ascommonly observed in species like Papilio polytes. Such limitationshave led to increasing reliance on molecular techniques, especially DNAbarcoding using the mitochondrial cytochrome c oxidase subunit I(mtCOX1) gene.This study focuses on identifying butterfly species in the biodiverseregion of Jharkhand, India, through integrative taxonomy combiningclassical morphology and molecular genetics. Butterfly samples werecollected from multiple sites, and DNA was extracted and amplifiedusing standard PCR protocols. The mtCOX1 gene was sequenced andanalyzed using bioinformatics tools such as BLAST, MEGA X, andClustalW. The resulting sequences were compared against globaldatabases (e.g., GenBank), and phylogenetic trees were constructedusing Neighbor-Joining, Maximum Likelihood, and Minimum Evolutionmethods.The molecular data successfully confirmed species identities, even inmorphologically ambiguous or mimetic forms, particularly in Papiliopolytes. Multiple sequence alignment and pairwise distance matricesshowed minimal intraspecific variation, supporting the robustness of mtCOX1 as a molecular marker. The study highlights the significance ofgene-based identification in resolving taxonomic ambiguities anddetecting cryptic diversity, which is often overlooked by morphologicaltaxonomy alone.In conclusion, this research demonstrates the efficacy of mtCOX1 genesequencing as a reliable, reproducible, and precise tool for butterflyspecies identification. It underscores the need for molecular techniquesto complement traditional taxonomy, especially in biodiversity-rich yetunderexplored regions like Jharkhand. The findings contribute toregional biodiversity documentation and set a foundation for futureconservation and ecological research initiatives.