Vector backbone integration in marker-free transformation events. Atlantic (H), Bintje (F) and Potae9 (W), were transformed with construct pBINAW2: Rpi-vnt1.1:Rpi-sto1. PCR analysis was performed using primers specific for tetA, trfA, NPTIII, ColE1, oriV and traJ to detect vector backbone integration. The plasmid pBINAW2:Rpi-vnt1.1:Rpi-sto1 was used as a positive control and the untransformed Atlantic as a negative control. Only the NPTIII primers amplified an a-specific fragment of similar size as shown here for the backbone free events W43-1 and W43-5 in untransformed Potae9. None of the other primers amplified an a-specific band in Potae9 or Bintje (data not shown) M: molecular weight marker.