The Awami League, a 14-party political alliance led by Sheikh Hasina, announced the blockade after Saturday's talks with Iajuddin Ahmed, the country's president and head of the interim government, broke down.
Ashim Kumar Ukil, an opposition official, said: "We are picketing 30 points in Dhaka to disconnect it from the rest of the country."
Thousands of Awami League supporters marched through the streets of the capital.
The blockade left the city deserted, with cars off the roads and many businesses and offices closed.
Around 12,000 extra police were stationed around the city.
The League is demanding the president's resignation as the head of a temporary body overseeing upcoming elections and a revision of a voters' list the US-based National Democratic Institute has said contains 10 million "errors" and duplications.
Without the reforms the opposition is concerned elections in January will be biased in favour of the outgoing government, led by the Bangladesh Nationalist Party (BNP).
The League accuses the BNP of trying to rig the elections by appointing party loyalists to key positions within the current caretaker administration and the election commission.
The interim-government said on Sunday that it will meet again soon with rival political leaders to discuss the issue.
At least 30 people have died, mostly in clashes between rival political activists, since the end of the BNP-led government's five-year mandate in October.
Several blockades have been imposed by the opposition in recent weeks, paralysing the nation and costing businesses millions of dollars.