After shooting an eight-under-par score of 280 on Thursday, round two was more of a struggle as the Tigers finished with a 295.
Duke, which was 10 shots behind Auburn after round one, shot a 10-under-par round on Friday to reel in the Tigers and take the lead with a 10-under-par 568 total.
Auburn, which is at 575, is one under par for the tournament and four strokes ahead of sixth place Vandy.
Senior Candace Schepperle shot the low round of the day for the Tigers with an even par 72. She is one under after 36 holes of play.
Auburn's Carlie Yadloczky putts on Friday at the Ironwood Country Club course.
Freshman Carlie Yadloczky shot a 72 to go with a 68 on Thursday and is Auburn's individual leader for the tournament and seventh overall.
Junior Cydney Clanton, who had an opening round 71, shot a 76 on Friday.
Patricia Sanz, who did not count in the scoring on day one with a 75, shot a 74 for the Tigers on Friday. Haley Wilson, who opened with a 70 for the Tigers, shot a 77 on Friday.
The top eight teams at each of the three NCAA regionals will advance to the NCAA Championships scheduled for May 18-21 in Wilmington, N.C.