President Barack Obama has said the US now formally recognises Syria's main opposition coalition as "the legitimate representative" of the Syrian people.
Mr Obama told ABC News that the National Coalition was now inclusive, reflective and representative enough for Washington to take this "big step".
The UK, France, Turkey and Gulf states have already given their recognition.
Russia said the US had decided to place all its bets on the coalition achieving an "armed victory".
The National Coalition for Syrian Revolutionary and Opposition Forces was formed last month and only a small number of countries have so far recognised it as the legitimate representative of Syrians.
Civil disobedience call
President Obama's statement came ahead of a meeting of foreign ministers from 70 countries to discuss the situation in Syria.
The meeting in Morocco is the first time that the Friends of Syria conference is attended by the Syrian National Coalition.
The leader of the National Coalition, cleric Moaz al-Khatib, addressed the meeting, calling on Syria's Alawite minority to rise up in a campaign of "civil disobedience" against the regime of President Bashar al-Assad, Reuters news agency reports.
Mr Assad's family are members of the Alawite sect and have used it as a key support base during their time in power. The opposition is dominated by Sunni Muslims, who form a majority of Syria's population.
Mr Khatib also said world powers, particularly Russia, would be held responsible if any attempt were made by Mr Assad's government to use chemical weapons.
The Friends of Syria group was due to acknowledge "the National Coalition as the legitimate representative of the Syrian people", according to a draft declaration.
The group includes major world powers, but not Russia and China - key backers of Mr Assad.
Activists say more than 40,000 people have died in the 18-month uprising against President Assad.
Reports from central Syria on Wednesday said a large number of civilians from the Alawite sect had been killed in a village to the west of Hama.
The circumstances are unclear but a video posted by opposition activists purported to show a young Alawite boy describing how government forces had destroyed a building where the civilians were being held hostage by pro-government militia.
The building, in the village of Aqrab, had been under siege from the Free Syrian Army and, according to the boy, as many as 300 civilians were killed,
There has been no word so far from the Syrian government and it is impossible to verify the activists' account.
More than half a million people have now fled the conflict in Syria to neighbouring countries, according to the UN's refugee agency.
Announcing the US decision to recognise Syria's main opposition, Mr Obama said it had earned the right to represent the Syrian people, but issued a note of caution as well.
"Obviously, with that recognition comes responsibilities," Mr Obama said.
"To make sure that they organise themselves effectively, that they are representative of all the parties, [and] that they commit themselves to a political transition that respects women's rights and minority rights."
Russian Foreign Minister Sergei Lavrov, expressing surprise at the announcement, said the US had decided "to place all its best on an armed victory of the Syrian National Coalition".
US recognition of the coalition does not mean the US will begin arming rebel groups, but officials told ABC that might be approved if it was thought to help achieve a political solution in Syria.
Addressing the Friends of Syria meeting, UK Foreign Secretary William Hague said on Wednesday that the UK did "not rule out any option to save lives".
"The Assad regime should not doubt our resolve, or miscalculate how we would react to any use of chemical or biological weapons against the Syrian people," he said.
Mr Obama warned that the US would not support extremist elements within the opposition coalition.
"Not everybody who's participating on the ground in fighting Assad are people who we are comfortable with," he said.
"There are some who, I think, have adopted an extremist agenda, an anti-US agenda, and we are going to make clear to distinguish between those elements."
One group he mentioned - the al-Nusra Front - is believed to be linked to al-Qaeda and responsible for nearly 600 violent attacks in major Syrian cities in the past year, the US state department estimates.
On Tuesday, US Secretary of State Hillary Clinton designated the group a foreign terrorist organisation, freezing any assets its members may have in US jurisdictions.