Objective To investigate the clinical application of fecal microbiota transplantation (FMT) in two kinds of colonoscopic cecal catheterization. Methods From January 2016 to June 2017, 200 patients who took colonoscopic cecal catheterization for fecal microbiota transplantation in our hospital were randomly divided into group A and group B, with 100 patients in each group.Group A used direct colonoscopy catheterization to complete catheterization, group B was treated by the second colonoscopy catheterization. The success rate of catheterization, catheterization time, average pain score and complication were compared between the two groups. Results Compared with group A, the time to reach the cecum in group B was slightly longer but not statistically significant (14.95min vs 15.26min,P=0.68). It had lower pain score (5.7 vs 4.8, P<0.05), lower complication rate. Conclusion Among the patients with colonoscopic cecal catheterization for fecal microbiota transplantation, there was no significant difference in the time of catheterization between the second colonoscopy and the direct colonoscopy, but it has the high success rate of catheterization and low pain, low risk, worthy of clinical promotion.