title: RNA-seq of T449 (autotetraploid rice) anthers,the anthers at single microspore stage, biological replicate 2
design: Total RNA was extracted from the autotetraploid rice (T449) anthers by using Trizol reagent (Invitrogen, CA, USA) following the manufacturer s procedure. The quantity and purity of total RNA were analysis by Bioanalyzer 2100 and RNA 6000 Nano LabChip Kit (Agilent, CA, USA) with RIN number > 7.0. Approximately 10  g of total RNA was used to deplete ribosomal RNA by using Epicentre Ribo-Zero Gold Kit (Illumina, San Diego, USA). Following purification, the poly(A)- or poly(A) + RNA fractions is fragmented into small pieces using divalent cations under elevated temperature. Then the cleaved RNA fragments were reverse-transcribed to construct the final cDNA library according to the manuscript of the mRNA-Seq sample preparation kit (Illumina, San Diego, USA). Finally, we performed the paired-end sequencing with the average insert size of 300 bp ( 50 bp) on an Illumina Hiseq2500 at the (LC Sciences, USA) following the vendor s recommended protocol.