It is just the second planned shuttle flight since the 2003 Columbia tragedy.
NASA will attempt to launch its seven astronauts on their key mission again on Tuesday -- US Independence Day -- at 2.38pm local time, to give launch crews a day of rest and to recharge fuel cells that power the shuttle.
The astronauts were once again sitting inside the space shuttle waiting for the green light when lift-off was cancelled two hours before its 3.26pm scheduled time.
"We had the vehicle ready to go, the crew ready to go and it's just that one thing that we really don't control, the weather, that kept us from launching,” said John Shannon, the mission management team's chairman.
Launch rules prevent NASA from sending Discovery up through the stormy clouds around Cape Canaveral.
"What a great gift NASA can give to the nation to return the shuttle to operation on Independence Day, so we are really looking forward to that," Mr Shannon told reporters at the Kennedy Space Centre in Cape Canaveral, Florida.
It would be the first July 4 shuttle launch.
NASA has until July 19 to launch Discovery, however if the orbiter does not get away by Wednesday, a hydrogen fuel cell system used to generate electricity on the shuttle will need to be replenished, pushing the launch even further back.
If the weather finally cooperates, the five men and two women astronauts will fly to the International Space Station (ISS) on a mission aimed at improving shuttle safety three years after Columbia disintegrated while returning to Earth.
During the 12-day flight, the astronauts will test new procedures to boost safety as well as deliver equipment and supplies to the ISS.
They will also drop off European Space Agency astronaut Thomas Reiter of Germany, who will join the ISS's two other crew members for a long-term stay, and perform two space walks.
NASA hopes to extend the mission by a day to conduct a third space walks to test new repair techniques.
The space agency has installed cameras on the shuttle and its fuel tank to detect any loss of foam during lift-off. The shuttle's heat shield will then be photographed as it approaches the ISS.
Columbia's demise was caused by a piece of foam that peeled off its external fuel tank during lift-off and pierced the shuttle's protective heat shield.
Debris also shed from Discovery's tank in last year's first post-disaster flight, but missed the shuttle. Still, NASA grounded the shuttle fleet until now to make further modifications to the troublesome fuel tank.
NASA Administrator Michael Griffin again defended his controversial decision to launch the spacecraft despite concerns expressed by his top safety official and chief engineer.
NASA chief safety officer Bryan O'Connor and chief engineer Chris Scolese had called for a six-month delay to the launch to redesign foam on the fuel tank.
But the two officials backed the launch after NASA said the astronauts could take refuge on the ISS and wait for a rescue mission should the shuttle suffer irreparable damage.