Abstract: In order to explore the effects of Longling Yellow goat-derived E. coli ClpV1 gene on some biological properties of E. coli, in the experiment, anal swabs of Longling Yellow goats with diarrhea were used as materials for bacterial isolation and identification; E. coli D1 strain（i.e. the original strain E. coli D1）, which was sensitive to both kanamycin and spectacularin, was selected from the isolates, and a deletion strain of the E. coli ClpV1 gene（i.e. the deletion strain E.coli ΔClpV1-D1） was constructed by using CRISPR/Cas9 technology. The genetic stability of the deletion strain E. coli ΔClpV1-D1 was tested by PCR after 15 consecutive generations. The original strain E. coli D1 and the deletion strain E. coli ΔClpV1-D1 were selected and inoculated in LB liquid medium, and their growth rates in the external environment were measured at 1 h intervals for 24 h. The original strain E. coli D1 and the deletion strain E. coli ΔClpV1-D1 were cultured at 30 ℃ for 72 h, and then the ability of biofilm formation was determined by crystal violet staining. Cell viability of the original strain E. coli D1 and the deletion strain E. coli ΔClpV1-D1 at 3, 6, 9, and 12 h was determined by MTT assay. The original strain E. coli D1 and the deletion strain E. coli ΔClpV1-D1 at concentrations of 1.0×10~5 cfu/mL and 1.0×10~8 cfu/mL were injected intraperitoneally into Kunming mice, respectively; the number of deaths in each group was recorded after 24 h, and the differences in pathogenicity were analyzed. The results showed that 12 strains of E. coli were isolated from 16 anal swabs; the deletion strain E. coli ΔClpV1-D1 was successfully constructed using CRISPR/Cas9 technology; the deletion strain E. coli ΔClpV1-D1 was still stable and inheritable even after 15 consecutive generations. There was no significant difference in growth rate between the original strain E. coli D1 and the deletion strain E. coli ΔClpV1-D1 at 24 h（P＞0.05）; the ability of the deletion strain E. coli ΔClpV1-D1 to form biofilm membranes was significantly reduced（P＜0.05）. The toxicity of the deletion strain E. coli ΔClpV1-D1 to IPEC-J2 cells was reduced in the four time periods measured, and the lethality of both concentrations in mice was significantly reduced（P＜0.05）. The results indicated that deletion of the ClpV1 gene had no significant effect on the growth rate of E. coli, but reduced its biofilm formation and pathogenicity.