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The MPs said in a letter to the speaker of the 360-seat lower chamber that they had joined the All Progressives Congress (APC) party.
This is the first time a president has lost his majority in the chamber since military rule ended in 1999.
Nigeria is due to hold presidential elections in 2015.
The defection of the MPs is the latest blow to Mr Jonathan and his governing People’s Democratic Party (PDP), and will make it extremely difficult for them to implement their legislative programme, correspondents say.
The PDP, which still controls the upper chamber, the Senate, has won every national election since the end of military rule.
Last week, ex-President Olusegun Obasanjo told Mr Jonathan it would be “morally flawed” for him to seek re-election because of his failure to tackle Nigeria’s myriad problems – including corruption, piracy, kidnapping and oil theft.
This came several weeks after a powerful faction of state governors broke away to join the APC.
The PDP now has fewer governors supporting it than the opposition.
The APC was formed in February following the merger of four opposition parties to challenge the PDP in the 2015 election.
Mr Jonathan moved from the vice-presidency to the presidency in 2010 after his predecessor, Umaru Yar’Adua, died in office.
He won presidential elections the following year.
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