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This article contains information on a subject that is not considered canon in the Gravity Falls universe.
Shriners temple, Atlanta, Georgia
"...The next clue, it's at Ochre Court. It's, uh, it's like, it's a big old building in Rhode Island. And if you go up the stairs, there's gonna be a bunch of pictures of nuns on the wall. You gotta look behind one of the nun pictures. Sister Mary Hilda Miley. Real, real grouch, that nun. Lift up the picture to find a clue behind the nun, all right? That's the whole thing..."
This pointed at Salve Regina University as the location of the next clue.
Salve Regina University, Newport, Rhode Island
The university accidentally disposed of the clue before the Cipher Hunt began, so Alex Hirsch promised to give a replacement clue at July 21, 2016, 12:30 PST, and asked people to stay out of the university in the meantime.
Century City, Los Angeles
The next time she made her appearance was on an audio file on a USB stick, which was used as a clue that was found in Century City, Los Angeles. This time she was heard as the Ghost of Seriousness, who hates everything fun and can only be defeated by "a clever solution," one of which being a vacuum cleaner, as heard when Soos used one on her. A transcript of that audio file can be found here.
She is a very strict nun who takes everything very seriously and hates fun things. Stan described her as a "real grouch". She beats people with a ruler.
Sister Mary has never been seen, but she is described by Stan as "some kind of ghost" who has "a ruler and a ghost hand."
"Mmmmm! It is me, Sister Mary Hilda Miley, and I decree that you shall never find the treasure! Muahahahaha!"
"I am the spirit of seriousness! I hate anything fun: carbonated beverages, boys and girls holding hands at school dances, and most of all, I hate treasure hunts! Also, knuckles!"
- Her name is based on a real person named Sister Mary Hilda Miley, who was associated with Salve Regina University.