title: NextSeq 500 sequencing; Stress Induced Calcium Signaling Facilitates ABA Mediated Transcriptional Adaptation
design: Stress Induced Calcium Signaling Facilitates ABA Mediated Transcriptional Adaptation
library_construction_protocol: Surface sterilized Arabidopsis thaliana seeds were sown on 0.5x Murashige and Skoog (MS) media (Duchefa, Netherlands) solidified with 0.8 % Agar (Duchefa, Netherlands) at pH 5.8 (adjusted with MES). After two days of stratification at 4 °C plates were transferred to growth chambers (Rumed, Rubarth Apparate GmbH) with a photoperiod of 16 h 22 °C: 8 h 18 °C, light : dark at 120-150 μmol m-2s-1 light intensity. Total RNA was extracted from approximately 75-100 mg mg frozen plant material with the SV Total RNA Isolation System (Promega Corporation, Madison, WI, USA). Quality of RNA was measured using an Agilent 2100 Bioanalyzer (Agilent Technologies).  rRNA was removed with the Ribo-Zero Plant rRNA removal kit for plant leafs (Illumina). RNA libraries were prepared using the NEBNext Ultra Directional RNA Library Prep Kit (New England Biolabs).