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• This was Mako's last film
• Oscar-nominated actor Mako was announced as the voice of Splinter at San Diego Comic Con the day before he died. Fortunately, he had finished the majority of his acting.
• Towards the end of the film, several items are shown that reference the live action movies. These include: Shredder's helmet and staff, A Foot Ninja Mask, the canister of ooze that mutated the turtles and Splinter (labeled TCRI instead of TGRI, though a crack is strategically placed to work either way), the Time Scepter, Walker's hat, and pieces of the battle suits of armor worn by the Turtles in the third film.
• During the first Nightwatcher scene, when the criminals are fleeing, one of the signs on the shops in the background spells out "monro" in hiragana, a Japanese writing system, which is most likely related to the writer and director Munroe.
• During Raphael's finishing lines, the phrase "we strike hard and fade away into the night" was used. That was also the ending line in the first TMNT comic.
• The film's lead track "Shell Shock" by Gym Class Heroes originated as a freestyle rap performed at a concert they performed at.
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