Gano was born in Connecticut to actor parents Norman and Faye Gano. The Gano family moved to Wisconsin in 1973, when his father opened an American Baptist church in Oak Creek, Wisconsin, a suburb of Milwaukee. Rev. Gano, an accomplished actor, also formed a community theater group in Oak Creek, and Gordon appeared in many of its productions, notably "Sing Out, Sweet Land" in 1976. His father played guitar, and exposed his son to a wide array of musical genres, including country and western, show tunes, and gospel. The liner notes to the Violent Femmes's compilation album Permanent Record describe Gano as "a devout Baptist".
In 1979 the Gano family moved to nearby Hales Corners, where his father took over an existing American Baptist congregation. At this time Gordon began attending Milwaukee public school—Rufus King High School, from which he graduated in 1981. After graduating from high school, he worked briefly as an encyclopedia salesman.
Older siblings from his parents' previous marriages living in the NYC area exposed the young Gano to such influences as The Velvet Underground, Lou Reed, Brian Eno, Patti Smith, Jonathan Richman, and The B-52's. Gordon's garage band days began with covers of Bob Dylan, The Beatles and moved through Grateful Dead (Hunter/Garcia) songs to Lou Reed and Velvet Underground songs by 1979. Gordon began writing his own songs in the mid '70s, influenced by the vocal stylings of Lou Reed, the story-telling of Hank Williams, Sr. and the poetics of Patti Smith's "Babelogue". Given his acting background, Gordon wrote many of his songs for characters, whom he would portray while performing the associated song.
Gano formed Violent Femmes in Milwaukee in 1980 with bassist Brian Ritchie and drummer Victor DeLorenzo, in which he was a guitarist, lead singer, and songwriter. They soon developed an enthusiastic following thanks to songs such as "Blister in the Sun", "Kiss Off" and "Add It Up" (all from their self-titled debut album). The band experimented with a variety of sounds over the course of their career, such as country and western (Hallowed Ground) and pop-rock (The Blind Leading the Naked).
During a Violent Femmes hiatus in the late 1980s, Gano formed a gospel-punk group called The Mercy Seat with vocalist Zena Von Heppinstall, bassist Patrice Moran, and drummer Fernando Menendez. They toured internationally for two years and released a self-titled album in 1987 on Warner Music Group's Slash Records. The album was re-released digitally in 2009 on Wounded Bird Records.
He released his first solo album in 2002, titled Hitting the Ground. He shared vocal duties (singing in Portuguese) with Manuel Cruz, the lead singer of the Portuguese rock band Ornatos Violeta in the song "Capitão Romance" for the last album released before their break-up, O Monstro Precisa de Amigos in 1999. Gano also played violin on Ben Vaughn's 1990 album Dressed in Black.
Gano contributed to the 2004 multi-artist collaboration The Late Great Daniel Johnston: Discovered Covered, alongside artists including Beck, The Eels, TV on the Radio, and Death Cab for Cutie, on which he provides a rendition of Johnston's "Impossible Love".
As of 2011 Gano is writing, recording and performing with ex-members of The Bogmen, Billy and Brendan Ryan, under the name Gordon Gano & The Ryans. Their first album, entitled Under the Sun, was released in September 2009 on Yep Roc Records.
He is also working with The Lost Bayou Ramblers on their upcoming release.
On September 14, 2013, Violent Femmes played at Chicago's Riot Fest, performing their entire first album, along with songs "Black Girls", "I Held Her in My Arms", and "American Music". Gano told the crowd of thousands that he had "forgotten how much fun doing this could be...thank you for reminding me."
Gano played Mr. Zank, the first of many substitute mathematics teachers, in an episode of The Adventures of Pete & Pete, entitled "X=Why?"
Gano is mentioned in the novel The Time Traveler's Wife by Audrey Niffenegger.
Gano appears as a fictional version of himself, under a love spell, in the 1997 Sabrina, the Teenage Witch episode "Hilda and Zelda: the Teenage Years"'.
The song "Good Feeling" is featured in the "Best Prom Ever" episode of How I Met Your Mother. The song is played at a high school prom by a band called The 88.