Arlene Foster has told Prime Minister Theresa May to focus on getting the right Brexit deal from Europe.
In a defiant message, the DUP leader warned against being pressured into accepting an unwise agreement which sold Northern Ireland short.
Writing in today's Belfast Telegraph, Mrs Foster repeats that she will not accept an Irish Sea border which "annexes" the region from the rest of the UK.
She said "many others" in Westminster and beyond supported her concerns.
It came after the DUP accused Mrs May of breaking promises over plans to avoid a hard border.
They were responding to a letter from the PM which the DUP interpreted to mean that she was planning a Brexit deal which could mean Northern Ireland becoming separated from the rest of the UK by a trade border in the Irish Sea.
In a day of dramatic developments:
Agreeing a backstop - the contingency plan to keep an open border until the UK and EU settle their future relationship - remains the main obstacle to a Brexit deal.
The leaked letter from the PM, in reply to an earlier message from Mrs Foster and her deputy, Nigel Dodds, set out Mrs May's approach.
The DUP believes the wording of Mrs May's letter means Northern Ireland-only measures will be contained in the Brexit deal, despite Mrs May's insistence it will never come into effect.
DUP Brexit spokesman Sammy Wilson warned his party - whose 10 MPs prop up Mrs May's government - would vote down the deal if it comes before the House of Commons.
Mrs Foster writes: "The desire to produce the right deal can be superseded by the perceived need to produce a deal."
Outlining her position on an Irish Sea border, she adds: "If what is outlined in the reply is the type of deal the Prime Minister intends to conclude, then the DUP could not support a deal which annexes Northern Ireland from the rest of the United Kingdom." She warns that Mrs May will "handcuff the UK to the EU, with the EU holding the key".
A Downing Street spokesman repeated the PM's commitment to avoiding a hard border.
Former Attorney General Dominic Grieve - a leading Remain advocate - told the BBC he recognised the DUP's position.
He said: "Once you start saying that you are going to have a different regulatory regime, which could include tariffs, could include all sorts of regulatory elements, then you would essentially have to have a hard border in the Irish Sea - and I don't see that as compatible with Northern Ireland's status within the United Kingdom."
Ulster Unionist leader Robin Swann questioned what influence the DUP had with the government.
Ms O'Neill said the backstop was agreed between the UK Government and the EU 27 last December, adding: "That remains the bottom line in order to prevent a hard border and safeguard our political and economic stability now and for the future.
"There can be no hardening of the border on the island, no undermining of the Good Friday Agreement and no loss of citizens' rights.
"The consequences of Brexit are real and they will be lasting. The backstop must be also."
I and the leaders of the other four pro-Remain parties will be in London on Monday and meeting An Taoiseach Leo Varadkar on Wednesday when we will be sending a strong, united message that the backstop, as already agreed, must be delivered.