Hostility to his idea voiced at the LibDem conference in Dunfermline means the First Minister will not have enough support to pass a bill at Holyrood.

Mr Salmond’s flagship policy was to have a vote on breaking up the Union late next year. But the lack of LibDem support means that, even with backing from the Greens and Independent MSP Margo MacDonald, the SNP can muster only 50 votes for a referendum bill, 15 shy of a majority.

The SNP said it planned to introduce its bill regardless, as it was a manifesto promise and they wanted the opposition parties to justify why they would deny people a say on their future.

But Labour and the Conservatives said ministers would only be wasting public money if they went ahead and urged them to be realistic.

Many MSPs believe the SNP exercise is a charade, as Mr Salmond does not want a referendum in case he loses and fatally undermines his authority.

Murdo Fraser MSP, deputy leader of the Scottish Conservatives, said it was time to “kill the bill”.

He said: “Alex Salmond dumped local income tax when he knew he would lose the vote in parliament. He should learn that lesson.

“Rather than waste time, effort and public money pursuing the minority whim of independence, the SNP should get on with what it was elected to do. The SNP should stop obsessing at the extremes of the constitutional debate.”

And Pauline McNeill, for Labour, said: “It is becoming increasingly clear that the SNP cannot win a majority for a referendum on independence.

“This issue is an unnecessary distraction when everyone in the Scottish Parliament should be working together to protect jobs and get Scotland through the recession.”

The SNP plan to issue a white paper on a referendum on St Andrew’s Day, and a bill in the spring, and had wanted a referendum in late 2010.

They only ever had an outside chance of passing a bill, as the three unionist parties oppose it. Ministers privately acknowledged defeat was likely, but kept pushing the policy to avoid splits within their own party.

However, in recent weeks tensions emerged with the LibDems on the issue, suggesting a referendum might be possible after all.

At the LibDems’ UK conference last month, Kevin Lang, the Westminster candidate for Edinburgh North and Leith, and Scottish MEP George Lyon both argued in favour of a referendum.

The party’s Holyrood leader, Tavish Scott, ordered former minister Ross Finnie to review the policy, and party members discussed the issue in a closed session yesterday. Although no vote was taken, sources said there was “no enthusiasm” for Mr Salmond’s bill.

But the question of whether the LibDems would support and help draft a referendum in the next parliament remains open, and Mr Scott could use the issue as leverage in any coalition talks.

Alistair Carmichael MP, the LibDem shadow Scottish Secretary, said: “The party overwhelmingly rejected Alex Salmond’s referendum bill.

“There was genuine and widespread anger at the rigged question put forward by the SNP.

“The Liberal Democrats are a party that will remain at the heart of the debate on Scotland’s constitutional future. But on our terms.”

A spokesperson for the constitution minister Michael Russell insisted a bill would still be issued: “The opposition parties will not be able to dodge the issue when it comes before parliament. Any party which votes against the people choosing Scotland’s future will pay a heavy electoral price.

“A majority of voters support a referendum, and we are focused on translating that popular support into a parliamentary majority next year, including being entirely open to including a ‘more powers’ question, alongside the option of independence and equality for Scotland.”