DOWNING Street has signalled for the first time that a vote to leave the EU could raise the prospect of a second Scottish independence plebiscite.

The warning came as David Cameron took a public swipe at Tory MP and London Mayor Boris Johnson, a leading light in the Leave campaign.

In barbed exchanges between the two big Tory beasts, the Prime Minister implied that, unlike him, Mr Johnson had a personal agenda to use the referendum to further his political career, with Downing Street his ultimate goal.

The Prime Minister used his statement to MPs to dismiss the idea – reportedly floated by Mr Johnson – that a Leave vote could be a prelude to securing a better deal in a second referendum.

Instead Mr Cameron made clear that a vote for Brexit would be followed by withdrawal negotiations under Article 50 of the Lisbon Treaty – the so-far unused procedure for leaving the EU.

Mr Cameron said: “Sadly, I’ve known a number of couples who have begun divorce proceedings, but I don’t know any who have begun divorce proceedings in order to renew their marriage vows.”

Around 200 business leaders have echoed Mr Cameron's concerns that Brexit would put the UK economy at risk in a letter to The Times. The signatories include the chairmen and chief executives 36 FTSE 100 companies, including Asda, BT, Mark & Spencer and Vodafone.

The Commons statement came as currency markets, spooked by the prospect of Brexit, saw sterling hit $1.40, its lowest level against the US currency for almost seven years.

No 10 normally issues a blanket response when asked about the prospect of a second independence referendum, citing the previous remarks of Nicola Sturgeon and her predecessor Alex Salmond that the 2014 poll was for a generation or even a lifetime.

But when asked yesterday if Mr Cameron believed those campaigning for Britain to leave the EU fully appreciated that one of the consequences of Brexit could be a second Scottish independence referendum, Mr Cameron’s spokeswoman replied: “There are many serious implications for the UK leaving the EU; no one knows the extent of them. That’s what those who are advocating Leave need to weigh up carefully and answer.”

Asked if Mr Cameron thought a vote to leave EU could lead to a Scottish vote in favour of independence, she added: “The PM is firmly focused on convincing people up and down the country that the best future for the United Kingdom is in the European Union.”

During Commons exchanges, Angus Robertson, the SNP leader, described the overwhelming extent of the support among Scottish politicians for the UK to remain in the EU.

He said: “If we are forced out of the EU, I am certain that the public in Scotland will demand a referendum on Scottish independence and we will protect our place in Europe.”

Mr Cameron replied by stressing that he would make a “positive case based around Britain being stronger, safer and better off” in the EU and added: “In terms of the vote in Scotland, this is one UK vote.”

His comments came after Ms Sturgeon predicted a Brexit would “almost certainly” lead to calls for another independence referendum.

The possibility of Scots voting to become independent from the UK in that event has been raised by several senior voices, including Tony Blair, Sir John Major, Nick Clegg and William Hague.

In December, Alan Johnson, the ex-Labour Home Secretary who is leading his party’s In campaign, said: “I don’t think there are many people who doubt if Britain votes to come out of Europe, Scotland will vote to come out of Britain.”

With well over 100 out of 320 Tory MPs backing Brexit, including several frontbenchers, the idea of a comradely contest up to the June 23 evaporated when Mr Cameron took a thinly-veiled swipe at Mr Johnson and his ambitions.

He told MPs: “I am not standing for re-election; I have no other agenda than what is best for our country.”

The PM mocked the London mayor’s suggestion that a Leave vote could be used to negotiate a better deal ahead of a second referendum, saying that decision in June would be final; the process of decoupling would start “straightaway”.

The Tory leader said the choice was between an “even greater Britain” by staying in or a “leap into the dark” by exiting the EU.

A day after the First Minister advised him not to campaign in Scotland because of his negative appeal, the PM told SNP MPs he was “looking forward to taking this message to Scotland” and smiled as he said he wanted to make the case for being “better together”.