Education Secretary Michael Gove says he would vote to quit the European Union if a referendum was held tomorrow.
Gove said life outside the EU would be "perfectly tolerable".
It came as pressure mounted on David Cameron as members of his own party attack last week's Queen's Speech, demanding instant legislation to guarantee his pledge of an in/out referendum.
Renegade MP Nadine Dorries, now back in the Tory fold, called for a formal pact with Ukip, while Nigel Farage's party continued to soar in the polls.
David Cameron has promised the referendum in the next Parliament, once he has attempted to renegotiate Britain's relationship with Brussels.
But this is not enough for some Tory backbenchers, and up to 100 are thought to be ready to back the rebel amendment if it is called, while ministers have been told to abstain.
Last year, it was reported that Gove had told friends he would vote to exit from the EU if a vote was held tomorrow.
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Gove said life outside the EU would be 'perfectly tolerable'
On the BBC's Marr Show on Sunday morning, Gove was asked whether he still thought this way.
He replied: "Yes. I am not happy with our position in the European Union but my preference is for a change in Britain's relationship with the European Union."
Gove said his position was to "recognise that the current situation is no good, to say that life outside would be perfectly tolerable, we could contemplate it, there would be certain advantages, but the best deal for Europe and for Britain would be if Britain were to lead the change that Europe needs."
Asked how he would vote, Gove added: "I am going to abstain."
He said he believed that a referendum in the future was needed, it could not happen now because of the coalition.
On Sky's Murnaghan show, Former Defence Secretary Liam Fox confirmed he would back the amendement, but said it was "not a big deal" because the vote would not matter.
Home Secretary Theresa May declined to say how she would vote in a referendum tomorrow, and said she would also be abstaining if the rebel motion goes to a vote.
Writing in The Sun, Dorries said it was time to consider joint Conservative and Ukip candidates.
"I do see a possibility that in areas where UKIP do not have a stronghold and where they feel they can endorse a Right-wing 'sound' candidate, it is something they may consider," she wrote.
"Very many of us in the Conservative Party would much rather be in bed with UKIP than the Liberal Democrats and I know who I would choose between Clegg and Farage as Deputy Prime Minister."