Nematodes are the most diverse but most minor studied microorganisms found in soil, water, animals, or plants. Either beneficial or pathogenic, they significantly affect human and animal health, plant production and ultimately affect the environmental equilibrium. Knowledge of their taxonomy and biology are the main issues to answer the different challenges associated with these microorganisms. The classical morphology-based nematode taxonomy and biodiversity studies have proved insufficient to identify closely related taxa and have challenged most biologists. Several molecular approaches have been used to supplement morphological methods and solve these problems with markable success. The molecular techniques range from enzyme analysis, protein-based information to DNA sequence analysis. For several decades, efforts have been made to integrate molecular approaches with digital 3D image-capturing technology to improve the identification accuracy of such a taxonomically challenging group and communicate morphological data. This review presents various molecular techniques and provides examples of recent advances in these methods to identify free-living and plant-parasitic nematodes. •Nematodes have astonishing diversity and most of them are unidentified.•Classical taxonomy has played a great role in identification of nematodes but presently facing shortage of taxonomists.•Molecular methods such as biochemical and PCR based methods have covered nematode diversity to some extent.•Advanced sequencing is a complete solution and, helps to create open databases for nematode identifications.