Soybean Transcriptional Response to Salt Stress

The salt treatments on soybeans were performed as previously described [22 (link)]. The seeds of the cultivated soybean accession C08 were germinated in vermiculite saturated with water. One-week-old seedlings were transferred to a hydroponic system with half-strength Hoagland’s nutrient solution. After the opening of the first trifoliate, the seedlings were treated with half-strength Hoagland’s solution containing 0.9% NaCl for different lengths of time (0 h, 1 h, 2 h, 4 h, 24 h, 48 h and 72 h). The roots and leaves of the treated plants were harvested separately and frozen in liquid nitrogen for total RNA extraction. Three individual plants of the same treatment were pooled as one sample and two independent biological replicates were performed for gene expression analyses.
Quantitative reverse-transcription PCR was performed using the PrimerScript one step RT-PCR kit (TaKaRa Biotechnology Co., Ltd., Dalian, China) according to the manufacturer's instructions. The primers for qRT-PCR are listed in Table S4. Relative gene expression was calculated using the 2^−ΔΔCt method [60 (link)]. The data were normalized to the reference gene ELF1b [61 (link)]. The qRT-PCR reactions were performed with at least three replicates.