SSR markers were identified from the Gramene database (http://www.gramene.org). 
PCR was performed using a total volume of 20 µl, which contained 10 ng template DNA, 0.2 
µM each primer, 50 µM dNTPs, 0.5 U Taq polymerase, and 2 µl 10X buffer with 1.5 mM Mgcl2. 
Thirty-five cycles were carried out, with an initial 3-min period at 94°C followed by cycles of 40 s 
at 94°C, 40 s at 55°C and 1 min at 72°C, and a final 10-min period at 72°C. PCR products were 
separated on 3% agrose gel.