Walker has not been charged as a part of the investigation, which a federal and state judge have halted. Prosecutors are appealing those decisions.
The case against Walker was first revealed in a court filing written in December but made public on June 19. The day after, Burke said she would not make the allegations a focus of her campaign.
"I'm going to focus on getting my message out to the people of Wisconsin," she said in an interview with WISN-TV that was taped on June 20. "The type of governor I'd be, what I believe in, how I'm going to move Wisconsin's economy forward - so it's not going to be a focus of my campaign."
Burke's new 30-second ad, which was airing in the Milwaukee, Madison, Green Bay, Wausau, and La Crosse markets, is titled "June 20." It includes footage of television anchors talking about the allegations as well as a federal economic report released last week that showed Wisconsin ranked 37th in new job creation in 2013.
Burke's spot criticizes Walker for launching his own negative ad on June 20 against her and concludes with the narrator saying, "Scott Walker just isn't working for you."
Watch the ad:
Burke's spokesman, Joe Zepecki, said the ad isn't about the alleged criminal scheme, but instead about Walker's decision to launch his own negative ad the day after that news and the latest jobs report broke.
But Walker campaign spokesman Tom Evenson said the ad was "yet another desperate attempt to distract voters from her failed record as Jim Doyle's commerce secretary."
Burke is a former Trek Bicycle Corp. executive and was Commerce secretary under former governor Doyle for two years. Walker's campaign, both in its latest ad and throughout the campaign, has tied Burke to Doyle's policies, including tax increases and job losses that occurred during the Great Recession.
"This election represents a clear choice for the voters: Do you want to continue the progress made under Governor Walker or do you want to take Wisconsin back to square one with Mary Burke?" Evenson said.
Wisconsin Republican Party director Joe Fadness said Burke is distorting Walker's record of having been governor at a time when the state added 102,000 new private sector jobs, after losing 133,000 jobs over the previous four years under Doyle.
Fadness also called Burke a hypocrite for bringing up the investigation after saying it wouldn't be a focus of her campaign.
"Burke's team abruptly reversed course from her pledge to not campaign on the politically motivated effort that has already been shut down by two judges, further evidence that she wants to deflect from her dismal record of outsourcing Wisconsin jobs and raising taxes on hard-working families," Fadness said.
Also on Thursday, the attorney representing lead prosecutor Francis Schmitz issued a statement saying that no determination had been made about whether to charge Walker or anyone else with a crime. The document released last week outlined prosecutors' theories, but "they did not establish the existence of a crime," attorney Randall Crocker said.
At the time the investigation was halted, Walker himself was not a target and had not been issued a subpoena, Crocker said. However, court documents show that Walker's 2012 recall campaign and a host of conservative groups were under investigation.
Walker was campaigning Thursday in Madison; Burke was in western Wisconsin. She is widely viewed as the front runner for the Democratic nomination, but she does face a challenge from state Rep. Brett Hulsey, who is mounting a longshot campaign. Walker is unchallenged.