Israel's Foreign Ministry said its embassy in Amman delivered the protest. It said it took the incident extremely seriously and had also summoned Jordan's acting ambassador to Israel to convey its objection.
A photo published on the Jordanian website Jfranews showed Information Minister Jumana Ghuneimat stepping on a large image of the Israeli flag.
Footprints were also printed on the flag, which Israel said was displayed at a gathering of Jordan's engineering union. The incident occurred last Thursday at Jordan's Trade Union building.
Israel and Jordan signed a peace agreement in 1994, but relations have often been frosty amid differences over Israeli policies in Jerusalem, where Jordan is custodian over Muslim sites, and toward the Palestinians.
Ghuneimat was not answering her phone on Sunday and did not issue any public statements.
Majed Qatarneh, spokesman of Jordan's Foreign Ministry, confirmed that Israel had sought clarifications over the incident and the ministry was handling the issue through "diplomatic channels."
"The Israeli side was informed that the building is a private one and the minister entered it from the main entrance for an official meeting," he said. "We have emphasized that we respect the peace treaty with Israel."