title: Metschnikowia fructicola 277 with grapefruit peel after 24h
design: RNA extraction and cDNA synthesis   Total RNA from frozen yeast pellet was extracted using MasterPureTM Yeast RNA Purification Kit (Epicentre® Biotechnologies, Madison, WI, USA) according to the manufacturer`s instructions.  Quality and concentration of total RNA was analyzed by gel electrophoresis and a spectrophotometer ND-1000, respectively (NanoDrop, Wilmington, DE, USA). First-strand cDNA was synthesized with Verso cDNA Kit (Thermo Fisher Scientific, Epson, UK), from 1 µg of total RNA that had been pretreated with 1.5 units of RNase I (Epicentre® Biotechnologies, Madison, WI, USA).    RNA sequencing   RNA samples isolated from control yeast M.fructicola grown in media (Mf) and yeast – fruit interaction (Mf-fruit) after 24 h of incubation and yeast – pathogen (Mf-Pdig) after 24 h of incubation were further prepared for RNA sequencing extracted using MasterPureTM Yeast RNA Purification Kit as described above. cDNA preparation, cDNA library construction, and sequencing were performed at The Genome High-Throughput Sequencing Laboratory, Sackler Faculty of Medicine and George Wise Faculty of Life Sciences, The Tel Aviv University. According to the Illumina manufacturer's instructions, poly(A)+ RNA was purified from 20 µg of total RNA using oligo(dT) magnetic beads and  fragmented into short sequences in the presence of divalent cations at 94 °C for 5 min. The cleaved poly(A)+ RNA was transcribed and second-strand cDNA synthesis was performed. After the end-repair and ligation of adaptors, cDNA products were amplified by PCR and purified using the QIAquick PCR Purification Kit to create specific cDNA library. Cluster formation, primer hybridization, and singled-end 76cycle sequencing were performed on an Illumina Genome Analyzer IIx using proprietary reagents according to the manufacturer’s instructions.
library_name: sa82