Egg ovotransferrin (OVT) is considered as a functional food ingredient for its various bioactivities. The objective of this work was to explore the potential biological activity of OVT on the gut health.Both young (3-week old) and adult (8-week old) mice models were utilized in this research. Each group received a standard diet containing either OVT (experimental group) or distilled water (control group) for a 14-day period. Transcriptome and 16S rDNA sequencing analyses were applied to characterize the gene expression in colonic epithelial cells and gut microbiota composition. In the young groups, OVT suppressed the genes correlated with lipid metabolism and signal transduction. The regulated genes in the adult groups encompassed various biological processes, including lipid metabolism, signal transduction, endocrine system, and others. OVT increased the proportion of some beneficial bacteria largely, especially Akkermansia, and inhibited some harmful bacteria. Furthermore, OVT affected mucosal morphology positively via increasing the crypt depth. OVT also increased the expression of tight junction protein occludin by 3.0 and 5.2 folds in young and adult groups, respectively.OVT exhibited some beneficial effects on gut environment. These positive findings provided new insight into the understanding of OVT as an excellent functional ingredient. This article is protected by copyright. All rights reserved.