biostudies-literatureZou ZPNational Natural Science Foundation of ChinaNational Key Research and Development Program of China102254https://www.ebi.ac.uk/biostudies/studies/S-EPMC10154985biostudies-literatureUnknown4(2)Engineered microorganisms hold potential for disease diagnosis and treatment. Here, we present a protocol to engineer E. coli Nissle 1917 strain (iROBOT) using genome insertion and plasmid construction to diagnose, record, and ameliorate inflammatory bowel disease in mice. We describe steps for constructing and administering iROBOT, diagnosing and recording colitis, preparing samples, and analyzing fluorescence and base editing ratios of iROBOT. We detail a colitis ameliorating assay using the disease activity index, colon length, tissue pathological section, and cytokine analysis. For complete details of the use and execution of this protocol, please refer to Zou et al.¹.STAR protocolsProtocol for engineering E. coli Nissle 1917 to diagnose, record, and ameliorate inflammatory bowel disease in mice.PMC101549852020YFA090880022134003Zou ZPFan YHYe BCDu WDu YFang TTZhou YfalseProtocol for engineering E. coli Nissle 1917 to diagnose, record, and ameliorate inflammatory bowel disease in mice.Engineered microorganisms hold potential for disease diagnosis and treatment. Here, we present a protocol to engineer E. coli Nissle 1917 strain (iROBOT) using genome insertion and plasmid construction to diagnose, record, and ameliorate inflammatory bowel disease in mice. We describe steps for constructing and administering iROBOT, diagnosing and recording colitis, preparing samples, and analyzing fluorescence and base editing ratios of iROBOT. We detail a colitis ameliorating assay using the disease activity index, colon length, tissue pathological section, and cytokine analysis. For complete details of the use and execution of this protocol, please refer to Zou et al.¹.2023-01-01T00:00:00Z2023 Apr2025-04-19T15:34:23.25Z2024-11-09T22:40:55.286ZS-EPMC101549853711566610.1016/j.xpro.2023.102254