The aim of the present study was to explore the clinical efficacy of mesocolon rotation and reverse puncture in total laparoscopic sigmoid vaginoplasty (LSV). The clinical data of 11 patients (unmarried, n=4; married, n=7) with congenital absence of a vagina who underwent treatment of total laparoscopic sigmoid vaginoplasty at the First Affiliated Hospital of Xinjiang Medical University (Urumqi, China) between March 2013 and March 2016 were retrospectively analyzed. In all patients, the surgical method included sigmoid mesocolon rotation and reverse puncture. The vaginal depth, the first sexual intercourse time and female sexual function indexes were recorded. The average operation time was 187±19 min, the average intra-operative blood loss was 132±24 ml, the time to the first meal after surgery was 4.3±1.1 days, the average post-operative hospital stay was 7.5±1.2 days, the post-operative short-term complication rate was 36.3% and the time to the first sexual intercourse was 3.0±0.3 months. The post-operative follow-up results indicated that the anatomical standard was reached in all of the 11 patients. Among the seven married patients, five patients were satisfied with their sex lives after the operation. In terms of psychosexual desire, only one married patient felt no sexual arousal. The other married patients had good sexual relations, function and satisfaction. In conclusion, the application of mesocolon rotation and reverse puncture in total LSV is safe and feasible.