Patrizia Stefanoni examined DNA traces in the aftermath of the 2007 killing of Meredith Kercher in Perugia, but her work was criticized by independent experts appointed by the court.
In testimony spread over two days, Stefanoni has rejected accusations of shoddy forensics work.
Experts have alleged glaring errors in evidence gathering, which raised doubts about the attribution of DNA traces and possible contamination of the evidence.
Debate over DNA evidence has taken the center stage at the appeals trial, with genetic experts taking the stand in the frescoed courtroom in Perugia.
In the original trial, prosecutors maintained that Knox's DNA was found on the knife's handle and that Kercher's DNA was found on the blade.
Stefanoni said the knife, found at Sollecito's house, was tested in a lab six days after the investigators had analyzed a trace of Kercher's DNA - excluding contamination.
Knox and Raffaele Sollecito were convicted of sexually assaulting and killing Kercher in the apartment that Knox and the 21-year-old Briton shared while studying in Perugia. Knox was sentenced to 26 years in prison; Sollecito to 25. Both deny wrongdoing and have appealed the December 2009 verdict.
A verdict is expected by month's end.
The Associated Press contributed to this report.