Gaiix analyzer

Total RNA was extracted using the TRIzol reagent (Life Technologies, Grand Island, NY), followed by purification using the RNAeasy mini clean up kit (Qiagen, Valencia, CA). Tissue samples were extracted individually, RNA from leaf, pod and root was then pooled in equimolar amounts and submitted for sequencing. The quality and quantity of RNA were examined using an Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA). Complementary DNA libraries were prepared and bar-coded for each of these accessions at the National Center for Genome Resources. RNA sequencing was performed on a GAIIx Analyzer (Illumina, San Diego, CA) for the tetraploid, and on a HiSeq 2000 (Illumina, San Diego, CA) for diploids.

Tissue samples for RNA extraction were obtained from five seedlings pooled together per replicate. Two replicates were extracted separately using the TRIzol reagent and company recommended protocols (Life Technologies, Grand Island, NY). The total RNA samples from each replicate were quantified using Nano Drop and subsequently purified using the RNAeasy mini clean up kit (Qiagen, Valencia, CA). The quality and quantity of RNA were examined using an Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA). Subsequent cDNA library construction and bar-coding for each of the samples from the two genotypes under each temperature treatment was conducted at the National Center for Genome Resources. RNA sequencing was performed on a GAIIx Analyzer (Illumina, San Diego, CA).

Genome sequences for three ST93 strains (TPS3104, TPS3105, TPS3106) were obtained from an Illumina GAIIx analyzer using 100 bp paired-end chemistry with a mean fold coverage of 331×. Genome sequencing of the two laboratory-induced mutants JKD6159∆hla (TPS3265) and JKD6159_AraCr (TPS3268) was performed using Ion Torrent sequencing technology.