Abstract: To understand the molecular mechanism of Micropterus salmoides subjected to ammonia nitrogen stress, the HiSeq sequencing strategy of Illumina platform was used to analyze the gene expression profiles in the liver of M. salmoides at 12h ammonia nitrogen stress (12h) and 24h ammonia stress (24h). We obtained 111.66 Gb of valid data and assembled a total of 133486 unigenes with N50 value of 1000 bp. Comparative transcriptional analysis identified 2072 differentially expressed genes (DEGs) at two time points. Specifically, 1516 DEGs were obtained at 12h, consisting of 907 up-regulated and 609 down-regulated genes, while 556 DEGs were obtained at 24h, with 330 up-regulated and 226 down-regulated. Gene ontology analysis revealed that the number of DEGs enriched at 12h was significantly higher than that at 24h, but there were more DEGs related to “Oxidoreductase activity” at 24h. In addition, KEGG enrichment analysis indicated that the DEGs were mainly involved in oxidative stress, autophagy and apoptosis related pathways. Subsequently, ten DEGs were selected for qPCR validation, and their expression levels were consistent with the sequencing analysis results, demonstrating the reliability of the sequencing data. Notably, the oxidative stress related gene Hypoxia-inducible factor 1 α (HIF1 α), Growth arrest DNA damage inducible protein 45β (Gadd45 β), DNA damage inducible transcript 4 (DDIT4), autophagy related gene microtubule-associated protein light chain 3 (LC3) and apoptosis downstream related gene endoplasmic reticulum oxidoreduclin1α (Ero1 α) displayed significant upregulated throughout the entire process of ammonia nitrogen stress. In summary, ammonia nitrogen stress could cause oxidative stress response in the liver of M. salmoides, while the damaged cells were eliminated by autophagy and apoptosis when the body’s regulatory role is insufficient to maintain internal environmental stability. The genes and pathways identified in this study will facilitate further investigations into the molecular mechanisms behind the tolerance of M. salmoides to ammonia stress.