Novotna, who won the Wimbledon title in 1998 when she defeated France's Nathalie Tauziat in the final, died on Sunday in her native Czech Republic surrounded by her family, a WTA statement said.
She played in two other Wimbledon singles finals, losing to Steffi Graf in 1993 and to Martina Hingis in 1997.
The picture of a distraught Novotna being comforted by the Duchess of Kent after losing the 1993 final when she had led 4-1 in the final set is one of the memorable Wimbledon moments.
Wearing her distinctive headband, she won doubles titles at all four Grand Slam tournaments.
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Czech-born American great Martina Navratilova led a number of players -- both past and present -- expressing deep sorrow at Novotna's death.
"The tennis world is so sad about the passing of Jana Novotna...I am gutted and beyond words- Jana was a true friend and an amazing woman..." Navratilova tweeted.
Former US player and now tennis commentator Pam Shriver -- who along with Zina Garrison beat Novotna and Sukova to Olympic doubles gold in 1988 -- remembered her rival fondly.
Lucie Safarova, the 2015 French Open finalist, also paid an emotional tribute to her fallen compatriot.
"Still shocked about this terrible news..you will be always remembered as our legend amazing athlete and a great person!" Safarova wrote on her Instagram page alongside a picture of Novotna clutching the Wimbledon trophy.
Steve Simon, chief executive of the WTA, said: "Jana was an inspiration both on and off court to anyone who had the opportunity to know her.
Born on October 2, 1968, Novotna won a total of 24 singles tournaments and 76 doubles tournaments in her career that spanned from 1987 to 1999.
She reached a high of world number two and became the oldest first-time Grand Slam singles champion at 29 years and nine months -- a record since eclipsed by Francesca Schiavone and Flavia Pennetta.
Novotna also won the Fed Cup with the former Czechoslovakia in 1988, as well as two Olympic doubles silver medals, and a bronze in singles at the 1996 Games in Atlanta.
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