Gene Expression Profiling under Gravitational Stimuli

The plants were either straight or inclined at 35° for 30, 60, 120 or 180 min in the isotropic device. Eight independent biological replicates (individual plants) were produced for each condition. XU and XL cDNA samples were used to monitor the expression of the 153 candidate genes by real-time qPCR at high-throughput. The experiments were performed according to the Fluidigm gene expression protocol without modification using the Biomark platform (Fluidigm, USA) and the 96x96 IFC gene expression (integrated fluidic circuit). Relative quantification was performed using the 2^−∆∆Cq method developed by Pfaffl (2001) (link). Expression levels were related to the geometric mean of the 153 expression levels of the candidate genes proposed by the Bestkeeper reference research software (Vandesompele et al. 2002 (link); Pfaffl et al. 2004 (link)), but using a larger subset of genes. No significant Tukey post hoc HSD was found in the change in gene expression (P < 0.05, data not shown) between qPCR and microarray, which validated the microarray results.