In recognition of his 25th anniversary on The Tonight Show, Carson received a personal Peabody Award, the board saying he had "become an American institution, a household word, [and] the most widely quoted American." They also said they "felt the time had come to recognize the contributions that Johnny has made to television, to humor, and to America."

Carson joined the United States Navy on June 8, 1943, and received V-12 Navy College Training Program officer training at Columbia University and Millsaps College. Commissioned an ensign late in the war, Carson was assigned to the USS Pennsylvania in the Pacific. While in the Navy, Carson posted a 10–0 amateur boxing record, with most of his bouts fought on board the Pennsylvania. He was en route to the combat zone aboard a troop ship when the bombing of Hiroshima and Nagasaki ended the war. Carson served as a communications officer in charge of decoding encrypted messages. He said that the high point of his military career was performing a magic trick for United States Secretary of the Navy James V. Forrestal. In a conversation with Forrestal, the Secretary asked Carson if he planned to stay in the navy after the war. In response, Carson said no and told him he wanted to be a Magician. Forrestal asked him to perform, and Carson responded with a card trick. Carson made the discovery that he could entertain and amuse someone as cranky and sophisticated as Forrestal.

In 1948, Carson married Jody Wolcott. The marriage was volatile, with infidelities committed by both parties, and ended in divorce in 1963.

To take advantage of the educational opportunities from the Navy, Carson attended the University of Nebraska in Lincoln, where he joined Phi Gamma Delta fraternity and continued performing magic (then paid $25 per appearance). He majored in journalism with the intention of becoming a comedy Writer. Instead, he switched his major to speech and drama a few months later, because he wanted to become a radio performer. Carson's college thesis, titled "How to Write Comedian Jokes", was a compilation of taped skits and jokes from popular radio shows with Carson explaining the comedic technique in a voice-over. It allowed him to graduate in three years. Carson graduated with a Bachelor of Arts degree in Radio and Speech with a minor in Physics in 1949.

Carson began his broadcasting career in 1950 at WOW radio and television in Omaha, Nebraska. Carson soon hosted a morning television program called The Squirrel's Nest. One of his routines involved interviewing pigeons on the roof of the local courthouse that would report on the political corruption they had seen. Carson supplemented his income by serving as master of ceremonies at local church dinners, attended by some of the same politicians and civic Leaders whom he had lampooned on the radio.

The wife of one of the Omaha political figures Carson spoofed owned stock in a radio station in Los Angeles, and in 1951 referred Carson to her brother, who was influential in the emerging television market in southern California. Carson joined CBS-owned Los Angeles television station KNXT. In 1953, comic Red Skelton—a fan of Carson's "cult success" low-budget Sketch comedy show, Carson's Cellar (1951 to 1953) on KNXT—asked Carson to join his show as a Writer. In 1954, Skelton during rehearsal accidentally knocked himself unconscious an hour before his live show began, and Carson successfully filled in for him. In 1955, Jack Benny invited Carson to appear on one of his programs during the opening and closing segments. Carson imitated Benny and claimed that Benny had copied his gestures. Benny predicted that Carson would have a successful career as a Comedian.

NBC's Tonight was the late-night counterpart to its early-morning show Today. Originating in 1953 with host Steve Allen, Tonight was somewhat experimental at the time, as the only previous network late-night program was NBC's Broadway Open House which starred Jerry Lester and Dagmar. Tonight was successful, and when Allen moved on to primetime comedy-variety shows in 1956, Jack Paar replaced him as host of Tonight. Paar left the show in 1962.

Carson hosted several shows besides Carson's Cellar, including the game show Earn Your Vacation (1954) and the CBS variety show The Johnny Carson Show (1955–1956). He was a guest panelist on the original To Tell the Truth starting in 1960, later becoming a regular panelist from 1961 until 1962. After the prime time The Johnny Carson Show failed, he moved to New York City to host Who Do You Trust? (1957–1962), formerly known as Do You Trust Your Wife?. In 1958, he appeared as a guest star in an episode entitled "Do You Trust Your Wife" on NBC's short-lived variety show, The Polly Bergen Show. On Who Do You Trust?, Carson met his Future sidekick and straight man, Ed McMahon. Although he believed moving to daytime would hurt his career, Who Do You Trust? was a success. It was the first show where he could ad lib and interview guests, and because of Carson's on-camera wit, the show became "the hottest item on daytime television" during his five years at ABC.

Although his show was already successful by the end of the 1960s, it was during the 1970s that Carson became an American icon and remained so even after his retirement in 1992. He adopted a Casual, conversational approach with extensive interaction with guests, an approach pioneered by Arthur Godfrey and previous Tonight Show hosts Steve Allen and Jack Paar. Former late-night host and friend David Letterman had cited Carson's influence.

Although he continued to have doubts about his new job, Carson became the host of Tonight (later becoming The Tonight Show Starring Johnny Carson) on October 1, 1962. After a difficult first year, he overcame his fears. While Tonight, under its previous hosts had been successful, especially under Paar, Carson's version eventually did very well in the ratings. Billy Wilder said of Carson:

In 1966, Carson popularized Milton Bradley's game Twister when he played it with Actress Eva Gabor. Not widely known at the time, the game skyrocketed in popularity after the broadcast.

Veteran NBC actor Raymond Burr became angry over Carson's continuing fat jokes and he only appeared on The Tonight Show in 1968 and in 1976.

Carson was a heavy smoker for decades and, in the early days of his tenure on Tonight, often smoked on-camera. It was reported that as early as the mid-1970s, he would repeatedly say, "These things are killing me." His younger brother recalled that during their last conversation, Carson kept saying, "Those damn cigarettes."

From July 1971, Carson stopped hosting five shows per week. Instead, Mondays featured a guest host, leaving Carson to host the other four weeknights. Shows were videotaped in Burbank at 5:30 pm, fed from there to the Central and Eastern time zone stations via cross-country television line at 8:30 pm Pacific time (11:30 pm Eastern time), and later sent from Burbank to the Pacific time zone stations at 11:30 pm Pacific time. Since only two feeds originated from Burbank, Central time zone stations received the Eastern feed one hour earlier at 10:30 pm local time, and Mountain time stations received the Pacific time zone feed one hour later at 12:30 am local time.

At the Carson Tonight Show's 10th-anniversary party on September 30, 1972, Carson announced that former model Joanna Holland and he had been secretly married that afternoon, shocking his friends and associates. On March 8, 1983, Holland filed for divorce. The divorce case finally ended in 1985 with an 80-page settlement, Holland receiving $20 million in cash and property.

In December 1973, Carson joked on Tonight about an alleged shortage of toilet paper. Viewers believed the story and panic buying and hoarding ensued across the United States as consumers emptied stores, causing a real shortage that lasted for weeks. Stores and toilet paper manufacturers had to ration supplies until the panic ended. Carson apologized in January 1974 for the incident, which became what The New York Times called a "classic study" of how rumors spread.

Carson was shown on a 1978 segment of 60 Minutes practicing at home on a drum set given to him by close friend Buddy Rich, who was the jazz musician with the most appearances on The Tonight Show. Gore Vidal, another frequent Tonight Show guest and friend, wrote about Carson's personality in his 2006 memoir.

Carson was head of a group of Investors who purchased and operated two television stations. The first was KVVU-TV in Henderson, Nevada, an independent station serving Las Vegas, acquired by the Carson group in 1979. Shortly after buying the station, KVVU was rumored to be acquiring an NBC affiliation as then long-time affiliate KORK-TV was in the process of being replaced by KVBC (and now KSNV), but it never happened. Carson's second station, independent KNAT-TV in Albuquerque, New Mexico, was purchased in 1982. Unlike the Las Vegas operation, KNAT faced stiffer competition for top-quality, syndicated programming. Carson sold both of his stations in 1985 and 1986 with KVVU-TV (FOX 5) going to the James Meredith Corporation and KNAT being sold to Trinity Broadcasting Network.

Carson often made jokes at the expense of other celebrities. In 1980, Carson backed out of a deal to acquire the Aladdin Hotel & Casino in Las Vegas, and a competing group led by Wayne Newton successfully bought the property. According to Lawyer Henry Bushkin, Carson became annoyed that he was often portrayed by the media as having "lost" the deal and reacted by telling jokes on his show about Newton, who had spent a great deal of effort building a masculine image. This created something of a high-profile feud between Carson and Newton. Years later, Newton appeared on Larry King Live, declaring that "Johnny Carson is a mean-spirited human being. And there are people that he has hurt that people will never know about. And for some reason at some point, he decided to turn that kind of negative attention toward me. And I refused to have it." Newton has often told of personally confronting Carson; after the final straw, Newton barged into his office at the studio and threatened to beat him up unless the jokes stopped. They did.

In 1981, Carson created the John W. Carson Foundation, dedicated to supporting children, education, and health services. The foundation continues to support charitable causes.

In 1982, Carson was found to be driving his DeLorean while under the influence of alcohol. He pleaded nolo contendere to a misdemeanor charge and received a sentence of three years' probation. Carson was required to attend an alcohol program for drivers and was permitted to use his car only to drive to work and back, without transporting any persons or animals in his vehicle.

On June 20, 1987, Carson married Alexis Mass. The marriage lasted until his death in 2005.

Carson had three sons, Christopher, Cory, and Richard. All three sons were from his first marriage. Richard Carson died on June 21, 1991, when his car plunged down a steep embankment along a paved Service road off Highway 1 near Cayucos, California. Apparently, Richard had been taking photographs when the accident occurred. On the first Tonight Show after Ricky's death, Carson paid tribute to Ricky's photographic work by showing his nature slides, accompanied by Stevie Ray Vaughan on blues guitar playing "Riviera Paradise". In addition, the final image of the show, as well as some "More to Come" bumpers, of Carson's last show on May 22, 1992, featured a photo Richard had taken.

Carson retired from show Business on May 22, 1992, at age 66 when he stepped down as host of The Tonight Show. His farewell was a major media event, often emotional for Carson, his colleagues and the audiences, and stretched over several nights. In tribute to Carson and his enormous influence, several networks that had late-night variety talk shows "went dark" for the entire hour he did the last show. The Tonight Show finally won the Emmy for Outstanding Late-night Series after 13 tries later that year, buoyed by the penultimate broadcast which featured Johnny's final two guests, Robin Williams and Bette Midler.

At the end of his final Tonight Show episode, Carson indicated that he might, if so inspired, return with a new project. Instead, he chose to go into full retirement, rarely giving interviews and declining to participate in NBC's 75th-anniversary celebrations. He made an occasional cameo appearance, including voicing himself on the May 13, 1993, episode of The Simpsons ("Krusty Gets Kancelled"), telephoning David Letterman on a November 1993 episode of Late Show with David Letterman, and appearing in the 1993 NBC special Bob Hope: The First 90 Years.

On May 13, 1994, Carson appeared on the Late Show with David Letterman. During a week of shows from Los Angeles, Letterman was having Larry "Bud" Melman (Calvert DeForest) deliver his "Top Ten Lists" under the guise that a famous personality would be delivering the list, instead. On the last show of the week, Letterman indicated that Carson would be delivering the list. Instead, DeForest delivered the list, insulted the audience (in keeping with the gag), and walked off to polite applause. Letterman then indicated that the card he was given did not have the proper list on it and asked that the "real" list be brought out. On that cue, the real Carson emerged from behind the curtain (as Letterman's band played "Johnny's Theme"), an appearance that prompted a standing ovation from the audience. Carson then asked to sit behind Letterman's desk; Letterman obliged, as the audience continued to cheer and applaud. After some moments, Carson departed from the show without having spoken to the audience. He later cited acute laryngitis as the reason for his silence. This turned out to be Carson's last television appearance.

Carson, an amateur Astronomer, was a close friend of Astronomer Carl Sagan, who often appeared on The Tonight Show. The unique way Sagan had of saying certain words, like "billions" of stars, would lead Carson to ribbing his friend, saying "BILL-ions and BILL-ions". Carson was the first person to contact Sagan's wife Ann Druyan with condolences when the scientist died in 1996. He owned several telescopes, including a top-of-the-line unit. In 1981, the minor planet 1981 EM₄ was named in his honor, 3252 Johnny.

On March 19, 1999, Carson suffered a severe heart attack at his home in Malibu, California, and was hospitalized in nearby Santa Monica, where he underwent quadruple-bypass surgery.

The 2005 film The Aristocrats was dedicated to Carson.

Carson reportedly loathed what he perceived as disloyalty, and he was furious when former Tonight Show guest hosts John Davidson and Joan Rivers began hosting their own talk shows. Rivers' show on the Fox Network directly competed with Carson during the 1986-1987 season before being cancelled. On June 24, 2009, following Ed McMahon's death, Rivers lauded McMahon on Larry King Live, but said that after she got her own show, Carson never spoke to her again.

In August 2010, the charitable foundation created by Johnny Carson reported receiving $156 million from a personal trust established by the entertainer years prior to his January 2005 death. Carson's foundation was now by far the largest of the Hollywood charities.

In November 2004, Carson announced a $5.3 million gift to the University of Nebraska Foundation to support the Hixson–Lied College of Fine and Performing Arts Department of Theater Arts, which created the Johnny Carson School of Theater and Film. Another $5 million donation was announced by the estate of Carson to the University of Nebraska following his death, while a $1 million donation was announced on November 4, 2011, creating the Johnny Carson Opportunity Scholarship Fund.

A two-hour documentary about his life, Johnny Carson: King of Late Night, aired on PBS on May 14, 2012, as part of their American Masters series. It is narrated by Kevin Spacey and features interviews with many of Carson's family, fellow comedians, and protégés.

Carson's other Business ventures included the successful Johnny Carson Apparel, Inc.—his turtlenecks became a fashion trend—and a failed restaurant franchise.

His lavish Malibu beachfront residence, valued at $81 million in 2017, contained only one bedroom. Friends and family members staying over would sleep in the guest house across the street.