Gloucester followed up last week's thrilling win over high-flying Exeter with another well-earned success as they cemented their chances of a play-off spot.
Ben Morgan scored two tries and Ed Slater one with Twelvetrees adding three penalties and three conversions.
Michael Rhodes, Jackson Wray and Matt Gallagher scored second-half tries in response for Saracens with Alex Lozowski kicking a penalty and three conversions. However, the Londoners look likely to lose their place at the top of the Gallagher Premiership table as second-placed Exeter face bottom club Newcastle at home on Saturday.
Gloucester had a good chance to take a second-minute lead but Tom Marshall's fumble prevented him from scoring as the wing was forced into touch a metre short of the line.
Minutes later, the hosts had another opportunity to go ahead, which again was squandered as Twelvetrees fired wide with a 40-metre penalty.
However, Gloucester continued to dominate the opening exchanges and were rewarded when Twelvetrees was on target with his second attempt.
Saracens then had their first chance for points but Lozowski missed a straightforward penalty so the home side led 3-0 at the end of an evenly contested first quarter.
Apart from Marshall's early miss the try-line had not been threatened until Gloucester produced a sweeping three-quarter move in the 21st minute which saw Ollie Thorley brought down in the opposition 22. But the pressure was maintained and gave Morgan the chance to force his way over for the game's first try.
Twelvetrees converted and added two penalties to one from Lozowski as Gloucester built up a healthy 16-3 advantage at half-time.
The home side made a mess of the restart to give Saracens early pressure to start the second half but much of their passing continued to be inaccurate, preventing them from gaining any momentum in the face of some stern defence.
Gloucester always looked more threatening in attack and it came as no surprise when they scored a second try.
Darts from Danny Cipriani and Willi Heinz put Saracens on the back foot and Slater was able to force his way over after Marshall was dragged down inches short.
The visitors rang the changes by substituting their whole front row and skipper Brad Barritt, who limped off to be replaced by Nick Tompkins.
Richard Wigglesworth also departed for Ben Spencer and there was an immediate dividend when Rhodes finished off their first real period of pressure, but Morgan's second try six minutes from time sealed their fate.
Saracens did manage to give the scoreline a more flattering look with late converted tries by Gallagher and Wray but the damage had already been done.
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