The key role of bioinformatics is acquiring striking importance in the era of outstanding advances in omics technologies for its fundamental support in describing the multifaceted aspects of biological functionalities. The manifold omics efforts flourishing worldwide are also contributing fundamental novelties in many aspects of agricultural sciences and, as a consequence, bioinformatics is acquiring a crucial role also in these research fields. Indeed, the transformation of natural environment for improvement of goods from animal, plants, and microbial worlds for human nutrition and health requires the comprehension of the molecular mechanisms influencing the structure and the function of the individuals, the populations, and the communities. The expanding knowledge about the molecules and the mechanisms associated with specific phenotypic traits and specific responses to biotic or abiotic stresses, complemented with the predictive power of bioinformatics, has an impact on agriculture practices and favors innovative methods in diagnostics, monitoring, and traceability, improving human benefits at lower costs, thus supporting sustainability. We here describe main bioinformatics approaches in the era of Next-Generation Sequencing for its impact in genomics, transcriptomics, and metagenomics efforts, describing their role in agriculture sciences. We aim to introduce common aspects, open questions and perspectives in this cutting-edge field of research.