One strain named as ETEC F4ac-GFP expressing green fluorescent protein（GFP）was obtained through transforming prokaryotic expression vector pTurboGFP-B into enterotoxigenic Escherichia coli（ETEC）F4ac by electrotransformation. It was identified that，by polymerase chain reaction（PCR），the obtained strain not only expressed the gene coding GFP，but also sent out green fluorescence as observed under a blue ray or a fluorescence microscope. The adhesion capacity of ETEC F4ac-GFP to IPEC-J2 cell lines was tested by in vitro adhesion test to verify the stability and reliability of its fluorescent expression at cell level，and the adhesion capacity to host cells was not changed when fluorescent plasmids were input. It was observed that，by cell counting kit-8（CCK-8）and real-time fluorescence quantitative PCR（qRT-PCR），the virulence of ETEC F4ac to host cells was not changed when fluorescent plasmids were input. A strain was thus provided as a tool for further researches on the pathogenesis of ETEC F4ac.