Nipponbare was grown on full nutrition for three
weeks. A total of 50 mg homogenized ground roots
and stems were used to extract RNA by using 10 mL
Trizol (Invitrogen) and RNA was further purified with
chloroform. After precipitation with 70% ethanol,
RNA was recovered with RNAEasy Mini Kit column
(Qiagen) and DNA was removed using the DNAase I
Kit (Qiagen) according to the manufacturer’s instructions
Semi-quantitative primers were designed by the
software Primer Premier 5 (PREMIER Biosoft). The
primer sequences were RTOsMAX1a (F/R), TCATC
TGGCCAGGCTCC/CCTGGGGTTCCCTTCATTGA
GC and RTOsMAX1e (F/R), ACCCCCTGGGGAG
ACTGCAT/CTCGATGCCGAGACGCCTCT.
cDNA was synthetized from 1 ȝg total RNA per
sample using the Script cDNA Synthesis Kit (BioRad)
following the manufacturer’s instructions. Finally, the
products were analyzed with 1.5% agarose gel
electrophoresis.