MPs have rejected plans to examine the PM’s Brexit bill in three days, leaving the legislation “in limbo”.
The Commons supported the Withdrawal Agreement Bill earlier, but voted against the short timetable.
Earlier, the PM warned he would seek an election if MPs dismissed the plan and the EU granted an extension to 31 October Brexit deadline.
After the vote, he told the Commons he would “pause” the legislation until he had spoken to EU leaders.
A spokesman from the European Commission said: “[The Commission] takes note of tonight’s result and expects the UK government to inform us about the next steps.”
Mr Johnson told MPs he was “disappointed” they had “voted for delay”, and said the UK “now faced further uncertainty”.
But he said his policy remained that Brexit would go ahead at the end of the month, adding: “One way or another, we will leave the EU with this deal to which this House has just given its assent.”
Labour leader Jeremy Corbyn said Mr Johnson was “the author of his own misfortune” – but offered to enter discussions over a “sensible” timetable for his deal to go through Parliament.
But the SNP’s leader, Ian Blackford, said it was “another humiliating defeat” for the PM, and MPs had “spoken with a very clear voice to tell the PM he is not on”.
Liberal Democrat leader Jo Swinson called on Mr Johnson to “end the brinkmanship and replace it with some statesmanship” in order to secure an extension with the EU.
Brexit bill’s journey so far
Boris Johnson agreed his new plan with EU leaders last week, but has repeatedly pledged to leave the bloc by the end of October, with or without a deal.
The bill that would turn his plan into law – the Withdrawal Agreement Bill – was published on Monday evening, and he urged MPs to back a three-day timetable to push it through the Commons ahead of the Halloween deadline.
The PM told Parliament if it “decides to delay everything until January or possibly longer”, he would seek an election – but he did not say what the government would do if the EU offered a shorter extension.
MPs did approve the bill on its first hurdle through the Commons – called the second reading – by 329 votes to 299.
But in a vote straight after, they rejected the so-called programme motion by 322 votes to 308 after a number of MPs criticised the pace of the legislation.
What happens next?
Mr Johnson told the Commons: “I will speak to EU member states about their intentions [but] until they have reached a decision – until we reach a decision, I will say – we will pause this legislation.”
In the meantime, however, he said the government would “take the only responsible course and accelerate our preparations for a no deal outcome”.
The PM added: “Let me be clear. Our policy remains that we should not delay [and] that we should leave the EU on 31 October.”
Irish Taoiseach Leo Varadkar said he welcomed the backing of MPs for the deal, adding: “We will now await further developments from London and Brussels about next steps, including [the] timetable for the legislation and the need for an extension.”
But Leader of the House Jacob Rees-Mogg said it was now “very hard to see how it is possible” to get a deal through Parliament by the end of the month.
If an election were to be triggered this week, the earliest it could take place would be Thursday 28 November, as the law requires 25 days between an election being called in Parliament and polling day.
But Mr Johnson cannot force an election himself and would need the backing of Parliament.
MPs had been due to debate the bill over Wednesday and Thursday, but will now return to discussing the contents of the Queen’s Speech – which put forward the government’s domestic agenda for the new session of Parliament.
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