GUS Staining

The RKN-infected galls and non-infected roots were fixed in 90% acetone for 24 h at -30°C and stained in the GUS buffer with 0.5mg/ml 5-bromo-4-chloro-3-indolyl-β-D-glucuronidase (X-Gluc; Wako) for 1 h. The reaction was stopped with Carnoy’s solution (90% (v/v) methanol, 10% (v/v) acetic acid), and the roots were mounted in a chloral hydrate solution (8 g chloral hydrate, 2 ml water and 1 ml glycerol), observed using an Axio Imager M1 microscope (Carl Zeiss Microscopy) and photographed using a DP71 Digital camera (Olympus). GUS assays with M. javanica and H. schachtii were performed according to Cabrera et al. (2014).