Total RNA was isolated from rice leaves using Isogen (Wako Pure Chemicals) followed by further purification with the RNeasy mini kit (Qiagen, Valencia, CA, USA). First-strand cDNAs were synthesized from equal amounts of total RNA using a ReverTra Ace qPCR RT Master Mix with gDNA Remover Kit (Toyobo, Osaka, Japan) in a total volume of 10 μl, as described by the manufacturer. qRT-PCR was performed with the Thermal Cycler Dice TP800 system (Takara, Tokyo, Japan) using a Kapa SYBR FAST qPCR kit (Kapa Biosystems, Cape Town, South Africa) as described by the manufacturer. The primers used for qPCR were as follows: BSR1 5′-AGGTGAGGTTGCACTCTGCT-3′ and 5′-CCAAGAATCCACCAACTCGT-3′ as described Dubouzet et al. (2011) (link); those for Rubq1 were 5′-GGAGCTGCTGCTGTTCTAGG-3′ and 5′-TTCAGACACCATCAAACCAGA-3′ as an internal control, as described Jiang et al. (2010) (link). Transcript levels of BSR1 were normalized to the endogenous rice reference gene (Rubq1).