Objective To determine the specific targets for spleen?kidney yang?deficiency ulcerative colitis (UC). Methods Ninety?six Wistar rats were randomly divided into the model, high dose, middle dose, and low dose sulfasalazine (SASP) groups. Then, next?generation sequencing was performed on colon lesions from the blank and model groups. Real?time quantitative PCR was performed to detect the mRNA expression of screened chemokines. Results Compared with the model group, differentially expressed genes in the blank group were screened according to a q?value of 0. 05 and fold?change of 1. 5. According to Gene Ontology classification analysis, the differentially expressed genes mainly functioned in three pathways: biological process, cellular component, and molecular function. Kyoto Encyclopedia of Genes and Genomes enrichment analysis of the differentially expressed genes showed alterations in the chemokine signaling pathway, and real?time quantitative PCR confirmed that CXCL1, CXCL2, CXCR2, CXCL6, CCL7, CCL12, and CCL7were significantly unregulated. Thus, the expression of the above factors was consistent with sequencing, and levels of these factors were decreased following treatment. Conclusions CCL12 expression was significantly unregulated along with the chemokine signaling pathway involving CXCL1, CXCL2, CXCR2, CXCL6, CCL7, and CCL12 in spleen?kidney yang?deficiency UC. Thus, this panel can be used as an objective index of inflammation for UC mucosa. Warming the spleen and kidney with Lizhong decoction and Sishen pills can effectively downregulate the expression of these factors, which restrained inflammatory reactions and promoted the repair of injured colonic mucosa.