From 7 a.m. to 10 p.m., ballots for the choice of "Yes" or "No" would be cast at 32 polling stations across Scotland, Xinhua reported.
This is the first Scottish independence referendum in Britain's history and official figures showed that about 97 percent of those eligible to vote in Scotland signed up to vote in the referendum, which is set to be the biggest poll in Scotland.
The results are expected to be announced early Friday.
If a majority of Scots vote for independence, Scotland would become independent March 24, 2016 after a period of negotiations with the rest of Britain.
If the "No" campaign wins, British Prime Minister David Cameron, leader of the Conservative, Labor Party Leader Ed Miliband and Deputy Prime Minister Nick Clegg, leader of the Liberal Democratic Party, offered more powers for Scotland starting Sep 19 in a joint pledge published Tuesday.
In October 2012, Cameron and Scottish First Minister Alex Salmond, leader of the ruling Scottish National Party, signed the Edinburgh Agreement, allowing Scotland to hold an independence referendum in 2014.
It is also the third referendum for Scotland after the two previous ones held respectively in 1979 and 1997 on Scottish devolution.
The referendum in 1979 failed to gain the mandatory 40 percent of the electorate, while the latter succeeded with an overwhelming majority of voters backing devolution.
As a devolved legislature, the Scottish parliament was reconvened in 1999 with authorities over some limited areas of home affairs, and the parliament of Britain keeps "reserved" powers including the ability to amend the terms of reference of the Scottish parliament.