Transcriptional reprogramming of floral and pod gene expression by ozone
Tissue sampling for RNA was done during R2 (full bloom), and R4 (full pod).  At each stage the appropriate tissue was sampled (full open flowers at R2 and initiating pods at R4).  Sampling was done at the middle 3-4 nodes in order to avoid compensation and senescence effects on the lower and uppermost nodes. RNA was extracted from each tissue using RNeasy Plant Mini Kit (Qiagen Inc., Valencia, CA USA) according to the manufacturer’s protocol. RNA quantity was be determined with a spectrophotometer (Nanodrop 1000, Thermo Fischer Scientific).  RNA quality was determined by running samples on the Agilent Bioanalyzer (Agilent Life Sciences, Santa Clara, CA USA). Genomic DNA contamination was removed from RNA samples using DNA-free DNase treatment (Applied Biosystems/Ambion, Austin, TX, USA) according to the manufacturer’s protocol. Each tissue type was tagged during library preparation. cDNA libraries were made using Illumina TrueSeq total RNA library prep kit.  Sequencing was done at the Biotechnology Center using the Illumina Genome HiSeq 2000 platform (www.igb.illinois.edu/biotech/htdna/services-equipment/illumina).