| Fredric March | ... | Samuel Langhorne Clemens (Mark Twain) |
 | Alexis Smith | ... | Olivia Langdon Clemens |
| Donald Crisp | ... | J.B. Pond |
| Alan Hale | ... | Steve Gillis |
| C. Aubrey Smith | ... | Oxford Chancellor |
| John Carradine | ... | Bret Harte |
| William Henry | ... | Charles Langdon (as Bill Henry) |
| Robert Barrat | ... | Horace E. Bixby, Riverboat Captain |
| Walter Hampden | ... | Jervis Langdon |
| Joyce Reynolds | ... | Clara Clemens |
| Whitford Kane | ... | Joe Goodwin, Editor |
| Percy Kilbride | ... | Billings, Enterprise Typesetter |
| Nana Bryant | ... | Mrs. Langdon |
| Jackie Brown | ... | Samuel Clemens - age 12 |
| Dickie Jones | ... | Samuel Clemens - age 15 |
| Russell Gleason | ... | Orion Clemens, Editor Habbibal Journal |
| Joseph Crehan | ... | Riverboat Captain / Ulysses S. Grant |
| Douglas Wood | ... | William Dean Howells |
| rest of cast listed alphabetically: |
| Ernie Adams | ... | Pickpocket (uncredited) |
| Gloria Ann Crawford | ... | Susie Clemens as a Child (uncredited) |
| Hooper Atchley | ... | Secretary (uncredited) |
| Arthur Aylesworth | ... | Worried Buffalo Merchant (uncredited) |
| Lynn Baggett | ... | Susie Clemens (uncredited) |
| Leah Baird | ... | Bit Role (uncredited) |
| Diana Barrymore | ... | Undetermined Role (uncredited) (unconfirmed) |
| Willie Best | ... | George, Twain's Butler (uncredited) |
| Henry Blair | ... | Boy (uncredited) |
| Oliver Blake | ... | Miner (uncredited) |
| Monte Blue | ... | Captain of 'Queen of Dixie' (uncredited) |
| Bob Burns | ... | Passenger (uncredited) |
| Viola Callahan | ... | Bit Role (uncredited) |
| Burr Caruth | ... | Oliver W. Holmes (uncredited) |
| Wheaton Chambers | ... | Joker at Train Station (uncredited) |
| Davison Clark | ... | Henry Wadsworth Longfellow (uncredited) |
| Chester Conklin | ... | Frog Jump Contest Judge (uncredited) |
| Carol Coombs | ... | Clara as a Child (uncredited) |
| Lloyd Corrigan | ... | Town Citizen (uncredited) |
| Frank Dae | ... | Houghton (uncredited) |
| Frank Darien | ... | Toy Store Proprietor (uncredited) |
| Earle S. Dewey | ... | Southerner (uncredited) |
| Dudley Dickerson | ... | Black Man (uncredited) |
| Dennis Donnelly | ... | Baby (uncredited) |
| Roland Drew | ... | Editor (uncredited) |
| Bill Edwards | ... | Bit Role (uncredited) |
| Sarah Edwards | ... | Bit Role (uncredited) |
| Dick Elliott | ... | Spectator at Frog-Jumping Contest (uncredited) |
| Jim Farley | ... | Prospector (uncredited) |
| Ross Ford | ... | Hardy - Howell's Secretary (uncredited) |
| Willie Fung | ... | Chinese Man (uncredited) |
| Rosina Galli | ... | Italian Nurse (uncredited) |
| Jack Gardner | ... | Bit Role (uncredited) |
| Jack Gargan | ... | Bit Role (uncredited) |
| William Gould | ... | Ship's Bouncer (uncredited) |
| Mildred Gover | ... | Black Woman (uncredited) |
| Jessica Grayson | ... | Housekeeper (uncredited) |
| William Haade | ... | Steam Room Chief (uncredited) |
| Frank Hagney | ... | Crew Chief (uncredited) |
| Creighton Hale | ... | Man with Mule (uncredited) |
| Thurston Hall | ... | Politician (uncredited) |
| George Haywood | ... | Bit Role (uncredited) |
| Robert Herrick | ... | English Boy (uncredited) |
| Harry Hilliard | ... | John G. Whittier (uncredited) |
| Gene Holland | ... | Huckleberry Finn (uncredited) |
| Harry Holman | ... | Drunken Guest (uncredited) |
| Stuart Holmes | ... | Man at Reception (uncredited) |
| Robert Homans | ... | Bit Role (uncredited) |
| Olin Howland | ... | Riverboat Southerner (uncredited) |
| George Humbert | ... | Bit Role (uncredited) |
| Brandon Hurst | ... | Ralph Waldo Emerson (uncredited) |
| Charles Irwin | ... | British Reporter (uncredited) |
| Kay Johnson | ... | Jane Clemens (uncredited) |
| Payne B. Johnson | ... | Boy in Frog Jumping Scene (uncredited) |
| Lew Kelly | ... | Pilot House Guest (uncredited) |
| Fred Kelsey | ... | River Rafter (uncredited) |
| Bill Kennedy | ... | Bit Role (uncredited) |
| Victor Kilian | ... | Higgins (uncredited) |
| Richard Kipling | ... | Bit Role (uncredited) |
| Bobby Larson | ... | Boy (uncredited) |
| Peter Lawford | ... | Young Oxford Celebrant (uncredited) |
| Billy Lechner | ... | English Boy (uncredited) |
| George Lessey | ... | Henry H. Rogers, Financier (uncredited) |
| Charles Marsh | ... | Prospector (uncredited) |
| Frank Mayo | ... | Pilot House Guest (uncredited) |
| Charles McAvoy | ... | Prospector (uncredited) |
| Sam McDaniel | ... | Comet Watcher (uncredited) |
| Sammy McKim | ... | Cub Pilot (uncredited) |
| John 'Skins' Miller | ... | Cross-Eyed Man (uncredited) |
| Michael Miller | ... | Tom Sawyer (uncredited) |
| Jack Mower | ... | Pilot House Guest (uncredited) |
| Paul Newlan | ... | Boss Deck Hand (uncredited) |
| Frank O'Connor | ... | Carriage Driver (uncredited) |
| Pat O'Malley | ... | Captain's Aide (uncredited) |
| Bud Osborne | ... | Lee Griggs (uncredited) |
| Paul Panzer | ... | Pilot House Guest (uncredited) |
| Charles Peck | ... | English Boy (uncredited) |
| Frank Pharr | ... | Stagecoach Passenger (uncredited) |
| Francis Pierlot | ... | Paige - Typesetter Inventor (uncredited) |
| Lee Powell | ... | Cowboy (uncredited) |
| Lillian Randolph | ... | Black Woman (uncredited) |
| George Reed | ... | Comet Watcher (uncredited) |
| Frank Reicher | ... | Doctor in Elmira, N.Y. (uncredited) |
| Betty Roadman | ... | Bit Role (uncredited) |
| Christian Rub | ... | Jones - Printer (uncredited) |
| Charlene Salerno | ... | Jean as a Child (uncredited) |
| Cliff Saum | ... | Prospector (uncredited) |
| Francis Sayles | ... | Bit Role (uncredited) |
| Paul Scardon | ... | Rudyard Kipling (uncredited) |
| George Sherwood | ... | Prospector (uncredited) |
| Emmett Smith | ... | Repeater (uncredited) |
| Walter Soderling | ... | Telegrapher (uncredited) |
| Frederick Spencer | ... | Jim (uncredited) |
| Libby Taylor | ... | Black Woman (uncredited) |
| Joyce Tucker | ... | Jean (uncredited) |
| Harry Tyler | ... | Assistant Editor (uncredited) |
| Dorothy Vaughan | ... | Katie Leary, Twain's Maid (uncredited) |
| Sailor Vincent | ... | Shopkeeper (uncredited) |
| Charles Waldron | ... | Dr. Quintard, Twain's Personal Physician (uncredited) |
| Eddy Waller | ... | Southerner (uncredited) |
| Lee 'Lasses' White | ... | Yokel (uncredited) |
| Leo White | ... | Barber (uncredited) |
| Ernest Whitman | ... | Stoker (uncredited) |
| Frank Wilcox | ... | Judge Clemens, Samuel's Father (uncredited) |
| Lottie Williams | ... | Lady in Audience (uncredited) |
| Norman Willis | ... | Ben Lake (uncredited) |
| Joan Winfield | ... | Bit Role (uncredited) |
| Harry Woods | ... | Bixby's Depth Caller (uncredited) |
| Harry Worth | ... | Gambler (uncredited) |
company.