Adrenaline Makeover: Played straight and subverted; June Havens certainly Took a Level in Badass after significant bits of the plot, but up until the halfway point of the movie she got progressively more worn down as the action progressed.
Always Save the Girl: Subverted: June deliberately gets herself kidnapped so that she can get back together with Roy, assuming that he'll save her. He does, but he has no idea that she's there at the time, as he's around for a completely different reason. He does take a moment in the middle of a firefight to step out from cover and kiss her when she guilt talks him into it, though.
Angrish: June's reactions in much of the first half of the movie.
Anti-Hero: Roy Miller may have killed dozens of innocents in cold blood and stolen the perpetual energy battery; he definitely kills dozens of agents in cold(ish) blood and drags an unknown innocent into the fight with him, though it's fairly obvious that he has little to no choice about the matter.
"Sorry, reflex. Hit me again; I won't dodge this time."
Artistic License - Physics: Subverted, but not at first. The Zephyr is presented as a true perpetual motion machine: it will never run out of energy, ever. However, over the course of the film, the battery slowly builds up heat and eventually explodes, taking out the villain in his moment of triumph.
Badass Driver: both Roy and June. June manages to pull off some amazing moves going the wrong way on the highway while controlling a runaway car from the back seat reaching over the corpse of the former driver: Roy calls her a "natural."
Belligerent Sexual Tension: Lampshaded by June after she's had a truth serum injected into her body and admits to Roy that all their crazy adventures make her want to have sex with him. His reaction is priceless.
Dueling Movies: Came out around the same time as Killers and while their plots are not identical, they are similar enough that the general public noticed the oddity of their close release dates. Ultimately, Killers ended up being panned(11% on Rotten Tomatoes) and barely turned a profit at the box office, while Knight and Day received more moderate reviews and grossed $260+ million.
Lodged Blade Recycling: June accidentally impales an assassin with a knife, followed by the assassin pulling it out of himself to attack June with.
Made of Iron: The assassin on the train. Wow. Most of the stuff is just standard movie things, until he gets stabbed in the chest with a knife! The assassin also No Sells June whacking him in the head with a frying pan.
Neutral Female: June, but not during the action sequences; she's actually this during much of the plot, being very decidedly on one side or the other when action breaks out.
Nice Guy: Roy, even when thrown into walls by super-strong Assassins, is a soft-spoken gentleman who does not curse or use swear words once throughout the entire movie. In one of the heart-warmeyer moments of the film, Roy has left June a note saying he was happy to have met her, and a second, reminding her to 'Eat a good breakfast, June'. He'd left her an omelette.
Precision F-Strike: June gets one in an argument with Roy about his "warning" not to get on a plane not being very effective, she then says that a better warning would've been saying something like "June, if you get on this plane, you will fucking die!"