title: RNA Seq of Saccharum officinarum X Saccharum spontaneum hybrid F2: Stem
design: Sugarcane stem and leaf tissues flash frozen in liquid nitrogen were finely ground with mortar and pestle and total RNA was extracted using Isol-RNA Lysis Reagent (5 PRIME Inc.; Cat # 2302700) as per manufacturer instructions. Additional isopropanol cleanup step was used to remove contaminants and enhance the quality of the total RNA.  The RNA were tested for integrity by running on agarose gels, and were quantified using NanoDropŽ Spectrophotometer.  RNA-seq libraries were prepared using KAPA Stranded mRNA-Seq Kit (Kapa Biosystems, Inc., Cat # KK8420) following instructions outlined by the manufacturer. Two microgram of total RNA per library in a 0.5x scaled down reaction volumes was used and final elution was done in 8ľl. The targeted fragment size for each library was 200-300 nt. The concentrations of eluted RNA-Seq libraries were estimated using QubitŽ Fluorometer and libraries yielding < 10ng/ľl were redone. Paired end sequencing of the libraries were carried out on HiSeq 2000 sequencer (IlluminaŽ) at UIUC-SIB Joint Center for Genomics and Biotechnology at Fujian Agriculture and Forestry University (FAFU).