In the level Blown' the Dam, there is a long Stair way with barrels are bouncing down at the player. Duke will comment, "Where's a hammer when you need one?" on the way up. At the top, you will see a Pig and Duke will say, "Huh, I was expecting a Monkey." This is an obvious reference to Donkey Kong.

In the level 'the Hive,' you can find the dead body of Isaac Clarke from Dead Space. Once you find him, Duke will comment, "That's one dead space marine."

During the first fight with an Assault Commander boss, Duke says, "I'm going to kill you old style!" This is a reference to the Youtube video Ventrilo Harassment where a Ventrilo user is harassed by someone with a Duke Nukem soundboard. The voice actor for Duke Nukem, John St. John, has mentioned in interviews that he has seen the Ventrillo Harassment video, which probably lead to the line being in the game.

"It's awesome, especially the Ventrillo Harassment! I LOVE that fans have used my voice to create some very funny videos and soundboards."

At one point during the game you will be accompanied by a Private Jenkins. After he runs ahead of you, he will be killed by an alien spaceship, and Duke will say, "Dammit Leroy!" This is a reference to the popular YouTube video where Leroy Jenkins gets his clan killed in World of Warcraft after charging into a dungeon screaming his name.

Duke was originally going to have a sidekick in the form of a gay robot. This wasn't to make fun of gay people, but to add depth to Duke's character. The idea was scrapped during development because the robot had a tragic ending, and the developers felt that Duke was more about action than tragedy.

"'Sexuality is part of the Duke personality.' The head of Gearbox explained that the original idea behind the sidekick was 'to explore how Duke would relate to a peer that might have a different sexual orientation.'"

"Ultimately the robot was in a situation where for in order for victory against the aliens to be accomplished, the robot's inner workings - a nuclear generator - had to be sacrificed."

"Duke is all about having a good time. Do you want to pay $60 to feel like crap? No!"

In the chapter "Damn, It's late," you will have to sign a book for a kid. This book (entitled "Why I'm So Great") is the same book that Duke was being interviewed about in Duke Nukem II.

In Duke's Titty City there is a message "Jenny 867-5309" scribbled on the wall. It is a reference to the song "867 5309 (Jenny)" by Tommy Tutone.

In Duke's Titty City, when you look in the mirror in the bathroom you can see the word "MURDER" reflected off the wall (as REDRUM) in the back. This is a reference to the movie The Shining.

In chapter Titty City when Duke interacts with a phone, he hears "Tonight... You." This is a reference to Handbanana from Aqua Teen Hunger Force.

When in the chapter Duke's Titty City, on the desk in the Employees Only room where the security camera is, there is a never-ending spinning top. This is a reference to Inception.

When opening the door to Queen Bitch, Duke says "You make me wish I had three guns." This is a reference to Total Recall movie, where a women with three breasts says "you're going to wish you had three hands." Queen Bitch also has three breasts.

In the Duke Dome chapter before fighting the boss, Duke says "you are one ugly motherfucker." This is a reference to the movie 'Predator'.

Duke is offered power armor, but he refuses the armor saying "Power armor is for pussies." This is a reference to the Halo series. It should also be noted that Halo themed music is playing in the background during the scene.

While in the casino, you see the numbers 4, 8, 15, 16, 23, and 42. It's a reference to the numbers from LOST.

At the chapter "Damn! it's Late" there is a moment that you walk back stage and encounter a dialogue between an actor and a director. This is a reference to Christian's Bale infamous freak-out toward his director during a movie scene shooting.

In chapter 'Damn, I'm Good' you can defeat the boss Energy Leech with just 1 hit from the Ray Freeze gun.

Before going into the dam, after Duke talks to the General, Duke sings "America, fuck yeah!" This is a reference to Team America World Police.

When you are in the casino, at the point you turn tiny and come across a mouse, Duke will say, "Now that's what I call a rodent of unusual size." This is a reference to The Princess Bride.

In the chapter 'Las Vegas' Duke says "I'm from Las Vegas, and I say: kill'em all!" This is a reference to Starship Troopers.

There is a soldier cursing and complaining that he had to help his friend find his wife. This is a reference to Marcus Fenix from Gears of War.