Nicola Sturgeon has denied that she could hold a wildcat referendum on independence if the SNP wins a majority in Thursday's Holyrood election.
Ms Sturgeon was speaking as she came under attack from Scottish Conservative leader Douglas Ross in a televised BBC Scotland leaders' debate.
Mr Ross claimed that the SNP would hold an "illegal" referendum if the prime minister refused to give formal consent.
But Ms Sturgeon replied: "No we won't."
The SNP leader accused Mr Ross of spreading "smears and untruths" about her position.
SIGN UP FOR SCOTLAND ALERTS: Get extra updates on BBC election coverage
She added: "What I have said consistently all along - sometimes to criticism from people on my own side of the argument - is that I would not countenance an illegal referendum, not least because it would not deliver independence.
"I want Scotland in the fullness of time and in due course to become an independent country.
"I will be responsible about that and I will build, and ultimately I think win, the case for independence through patient persuasion of people across the country."
SCOTLAND'S ELECTION: THE BASICS
What's happening? On 6 May, people across Scotland will vote to elect 129 Members of the Scottish Parliament (MSPs). The party that wins the most seats will form the government. Find out more here.
What powers do they have? MSPs pass laws on aspects of life in Scotland such as health, education and transport - and have some powers over tax and welfare benefits.
The referendum in 2014 - which saw voters in Scotland reject independence by 55% to 45% - was formally approved by the UK government, with both it and the Scottish government pledging to respect the result.
Prime Minister Boris Johnson has so far refused to grant the same consent for another vote on the issue, arguing that the 2014 referendum had settled the question for a generation.
And some within the independence movement - including former SNP leader and first minister Alex Salmond, who now leads the Alba Party - have called on Ms Sturgeon to set out an alternative path to independence if the PM's stance does not change.
And there have been suggestions that a referendum could be held without the UK government's approval - similar to the disputed vote in Catalonia in 2017.
The SNP manifesto says it wants to hold a referendum after the pandemic ends - with Ms Sturgeon not ruling out going to court to settle the question of whether Holyrood could legislate for a legal referendum without Westminster's backing.
The striking thing about the early stages of this debate was that when you get away from the constitutional faultlines, these parties actually have a lot in common.
The leaders all broadly agreed about taking a cautious route out of the Covid-19 lockdown, and all seem to be planning a "staycation" this year rather than international travel.
All want to commit more money to health and care services - and none of them want to raise income taxes to do it.
They are still politicians, so there were digs aplenty, but there were also lots of "I agree with..." moments - even between Patrick Harvie and Douglas Ross, who are not usually exactly chummy.
So there is plenty of common ground between the parties - but it tends to be forgotten when the debate swings to independence, and that was predictably the section where the "no talking over each other" rule went out the window.
Read more about Phil's five key "takeaways" from Tuesday night's debate.
But Mr Ross claimed: "Nicola Sturgeon has been clear. If she gets a majority, she'll take her eye off the ball for Scotland's recovery, for rebuilding the country from the pandemic, and seek to hold another independence referendum.
"She will ask for a Section 30 order - which the prime minister has said is absolutely the wrong thing to do right now in the middle of this global pandemic - and seek to divide the country all over again."
He said that, when the prime minister refused her request, Ms Sturgeon would "go ahead with an illegal wildcat referendum" - pointing to previous comments made by SNP MP Richard Thomson, who said the Scottish government would "simply go ahead with a referendum" if formal consent is refused.
What are the parties promising you?
Use our concise manifesto guide to compare where the parties stand on key issues like Covid-19, independence and the environment.
The televised debate also featured the leaders of the three other parties who are represented in the Scottish Parliament.
Scottish Labour leader Anas Sarwar said Mr Ross was the "gift that keeps on giving" for the SNP and claimed he was "only interested in saving his skin, not saving the union".
And he said it was "not credible" for Ms Sturgeon to lead the country through the recovery from the pandemic, while also leading a referendum campaign.
Mr Sarwar added: "I do not support a referendum, I don't support independence and I want people to choose something different.
"I want them to choose us to focus on a national recovery in the next parliament."
Scottish Liberal Democrat leader Willie Rennie said people would be "bemused" that party leaders were debating independence in the wake of one of deepest pandemics the country has ever faced.
Mr Rennie said: "The people who are waiting an age for mental health treatment I think deserve better than this, the people who are desperate for a job deserve better than this."
And he later added: "Despite the fact that it has been the policy of the SNP for generations, they still don't know what the currency would be" in an independent Scotland.
But Scottish Greens co-leader Patrick Harvie said the possibility of an independent Scotland would allow the country to shape a more radical future.
He added: "I don't think we can afford to pass up the opportunity to shape our own recovery. This is a moment of incredible opportunity to decide what kind of country is going to emerge from Covid."
But Mr Harvie and Ms Sturgeon disagreed over what currency an independent Scotland should use - with the Greens co-leader saying plans for a separate Scottish currency should begin "immediately" after a vote for independence.
But Ms Sturgeon said Scotland would continue to use the pound "for as long as necessary" after independence.
The debate also saw the five party leaders face questions on topics including the possible easing of foreign travel restrictions, the future of the social care sector, taxation and plans for a new £200m Royal Yacht.