Narm Charm: The musical intro where the Joes take out Cobra on Liberty Island from blowing up the Statue of Liberty where the over-the-top action, the ridiculousness of Cobra's attack, and the cheesiness of the song ("Crashing through the sky! Comes the fearful cry of...Cobraaaaa! (Cobraaaaaaaaa!) Cobraaaaaa! (Cobraaaaaaaaaaa!)") makes it looks like The War on Terror: The Musical. It's also beautifully animated, high tempo action, and a perfect distillation of the cartoon series in a single action sequence.
Signature Scene: The opening sequence. G.I. Joe foils a Cobra attack on the Statue of Liberty, with almost every single character who had ever appeared on the show contributing. Throw in Awesome Music and great moments like Duke punching Cobra Commander off the Statue.
The part of the vine trap sequence where Quick Kick, Snake Eyes, and Lady Jaye fall victim may or may not skirt the line between Fanservice and Fan Disservice.