Technical Abstract: Cervical anatomy limits the use of transcervical intrauterine artificial insemination (TC AI) in sheep. We have developed an instrument to cope atraumatically with the cervix. Even though this instrument has not affected fertilization rate or pregnancy rate through Day 3, the effects on sperm transport and pregnancy after Day 3 are not known. The objective of the present study was to determine whether our TC AI instrument affected sperm transport, pregnancy rates, or lambing rate. In Exp. 1, ewes were assigned to two treatments: TC AI using the new TC AI instrument (n = 10) or AI via laparotomy using a laparoscopic AI instrument (n = 10). Twenty hours after artificial insemination, the uterine horns and oviducts were recovered and flushed to collect spermatozoa. Sperm transport did not differ between the two treatments (P > 0.05). In Exp. 2, ewes were assigned to three treatments: TC AI using the new TC AI instrument + sham intrauterine AI via laparotomy (n = 29); sham TC AI + intrauterine AI via laparotomy using a laparoscopic AI instrument (n = 29); and sham TC AI + intrauterine AI via laparotomy using the new TC AI instrument (n = 30). On Day 14 after AI, uteri were collected and flushed to recover blastocysts. Transcervical deposition of semen reduced (P < 0.05) Day 14 pregnancy rate (17.2 vs 61%), but intrauterine deposition of semen using the TC AI instrument via midventral laparotomy increased (P < 0.05) Day 14 pregnancy rate (76.6 vs 44.8%). In Exp. 3, ewes were assigned to two treatments: sham cervical manipulation (n = 40) or cervical manipulation to mimic TC AI (n = 40). Immediately after treatment, each ewe was mated with a ram and watched until the ram mounted and ejaculated into the ewe. Treatment did not affect Day 30 or 50 pregnancy rate (67.5 and 66.2%, respectively), determined ultrasonically, or lambing rate (62.5%). The differences between Day 30 and 50 pregnancy rates and lambing rate were not significant. In Exp. 4, ewes were assigned to two treatments: TC AI (n = 99) or laparoscopic AI (n = 99). Transcervical AI reduced (P < 0.01) Day 30 (TC AI vs laparoscopic AI; 5.0 vs 46.0%) and 50 pregnancy rates (4.0 vs 41.0%), determined ultrasonically, and lambing rate (4.0 vs 41.0%). Even though the TC AI procedure significantly reduced pregnancy and lambing rates, large numbers of spermatozoa deposited at natural insemination seemed to compensate. Because our TC AI procedure has all but eliminated any visual evidence of trauma, and because the procedure does not seem to affect sperm transport or embryonal survival until Day 3, we speculate that cervical manipulation associated with TC AI may activate pathways that interrupt pregnancy between Day 3 and Day 14.