Addressing business leaders in Dublin, the Deputy First Minister highlighted the benefits of belonging to the EU and warned Prime Minister David Cameron's decision to put a renegotiated membership deal to a public vote caused "con-siderable uncertainty" for firms. Pro-UK parties – including the Tories – argued the independence referendum was damaging the Scottish economy.
The SNP was also accused of placing the country's EU membership in doubt after Ireland's European affairs minister, Lucinda Creighton, warned an independent Scotland would have to apply for membership in a "lengthy process".
Ms Sturgeon, responsible for the independence referendum, spoke after Mr Cameron confirmed plans for a 2017 poll on EU membership, if the Tories win the 2015 General Election.
She told the British Irish Chamber of Commerce annual conference: "It is no longer fanciful to consider as real the possibility the UK is heading out of the EU. Let me state emphatically this is not a journey the Scottish Government wishes Scotland to be a part of, nor one I believe would be supported across the Scottish business community or wider civic society. But it is a position that brings with it considerable uncertainty and one that could damage Scotland's EU interests and influence, and deter foreign investors from investing in our economy. A misguided policy that threatens tens of thousands of jobs in Scotland."
Patricia Ferguson, the constitution spokeswoman for Scottish Labour – who oppose the EU referendum – said: "Nicola Sturgeon is showing incredible hypocrisy by claiming a referendum on Europe is causing uncertainty when she makes Scots wait three years to have their say on staying in the UK."
Scots Tory chief whip John Lamont said: "The SNP has a cheek to accuse anyone of creating uncertainty."
The Nationalists claimed only a Yes vote in 2014 could guarantee Scotland's membership of the EU. However, opposition parties claimed Scotland's future membership was unclear under SNP plans after Ms Creighton said a newly independent country would face a lengthy negotiation process.
She said the application –which would have to be ratified by 27 member countries – might not take as long as Iceland, which remains an EU candidate state. It requested membership four years ago.
However, she added: "I don't see why it would be a terribly complex process."