GSM9076894: UBQ_33A_NULL_Rep01; Glycine max; RNA-Seq
Total RNA was isolated from ground frozen tissue with RNeasy (Qiagen Inc.), according to the manufacturer's protocol. Total RNA was then analyzed for quality and quantity with either the Bioanalyzer RNA Nano kit (Agilent Technologies) or the Fragment Analyzer with RNA  kit (Agilent Technologies) and normalized to 1 μg input per sample. Sequencing libraries were prepared using the Illumina Stranded mRNA library kit, according to vendor protocol (Illumina Inc. San Diego, CA). Briefly, mRNAs were isolated via attachment to oligo(dT) beads, fragmented, and reverse transcribed into cDNA by random hexamer primers with Superscript II reverse transcriptase (Life Technologies). The resulting cDNAs were end repaired, 3′-A-tailed, and ligated with Illumina indexed TruSeq adapters. Ligated cDNA fragments were PCR-amplified with Illumina RNA/DNA UDI indexed-primers, purified with AmpureXP Beads (Beckman Coulter Genomics), and checked for quality and quantity with the Agilent TapeStation 4200 system with D1000 ScreenTape. Libraries were combined into one sequencing pool based on equal-nm pooling, and pool then diluted to 2 nM. The pool was denatured according to Illumina sequencing protocols, hybridized, and clustered on an appropriate Illumina flowcell targeting 20M reads per sample.   Samples were sequenced using Single-Read 100bp reads.