Quantitative Gene Expression Analysis in Jatropha

Total RNA was extracted from each tissue, and first-strand cDNA was synthesized with a PrimeScript^® RT Reagent Kit with gDNA Eraser (Takara, Dalian, China) according to the manufacturer’s instructions. qRT-PCR was performed with LightCycler^® 480 SYBR Green I Master (Roche, Indianapolis, IN, USA) on the Roche 480 Real-Time PCR Detection System (Roche Diagnostics, Mannheim, Germany). qRT-PCR was performed with two independent biological replicates (tissue samples were harvested from different plants) and three technical replicates for each sample. Data were analyzed using the 2^−ΔΔCT method as described by Livak & Schmittgen (2001) (link). Expression levels of specific genes were normalized to that of the actin gene in Jatropha (Zhang et al., 2013 (link)). Primers used in qRT-PCR are listed in Table S3.