<p>The endoplasmic reticulum is a considerable network of intracellular networks. Its major functions are protein folding, proteosynthesis, post-transcriptional modifications, lipid anabolism, and sorting of the protein. ER isolation is very challenging because its biochemical properties are mostly like the other cellular endomembrane. Most previously published protocols for isolation are based on the density gradient ultracentrifugation. In this comprehensive study, we aim to generate data on the ER proteins of a chickpea plant and provide a reference map of proteomics analysis. Through the Easy-nlc LC–MS/MS technique, we identified 2192 proteins, including 338, and 681 unique proteins in sample 1 and 2, respectively, comprising both anticipated and newly discovered constituents. These identifications of proteins were validated through sequence analysis. In terms of biological significance, pulses play a major contribution to sustainable agriculture by enriching the soil with N₂ while maintaining a low water potential. One of the most cultivated legumes chickpeas is an essential source of protein and energy for both humans and animals. As the ER is responsible for protein folding and maturation, their proteome enhances our understanding of plant function. Here in this article, we optimized the isolation protocol from chickpea tissue through an ultracentrifugation process. Combined with the enzymatic analysis and proteomics analysis, these data will facilitate our better understanding towards the ER proteome study in chickpea plants.</p> Graphical Abstract <p></p>