Obasanjo ordered the probe of himself and other top officials after Orji Kalu, governor of southeastern Abia state, accused them of receiving bribes for oil contracts and privatisations, and operating illegal foreign bank accounts.
"The governor wrote a letter to the president making serious allegations against the president, senators and senior government officials, accusing them of corruption," said a spokesman for the Economic and Financial Crimes Commission
"On receipt of this letter, Mr President ordered the EFCC to investigate and make their findings public."
Kalu, a vocal critic of Obasanjo in the ruling People's Democratic Party (PDP) who has already announced his intention to run for president in 2007, accused senators and presidential aides of taking large bribes in return for government contracts in the oil and defence industries.
Saying he had "overwhelming evidence", Kalu accused Obasanjo of operating an illegal foreign bank account, and accused officials of taking bribes in the recent privatisation of two steel mills at Ajaokuta and Aladja.
He also accused Obasanjo of using public funds five years ago to build annexes to a university and secondary school allegedly owned by the president.
"Once an allegation is made, it must be investigated," said EFCC chairman Nuhu Ribadu.
It was the second high profile accusation of corruption against Obasanjo since he stepped up a campaign against graft earlier this year, sacking his education minister and police chief for suspected bribery and embezzlement.
A lawyer accused Obasanjo in May of corruption and abuse of power for raising 4 billion naira ($30 million) towards building a personal library in his home state.