title: plant sample from Cucumis sativus infected with CGMMV rep3
design: Total RNA was extracted from CGMMV-infected and not infected cucumber fruit peel, using the GenElute Mammalian Total RNA Miniprep Kit (Sigma-Aldrich) according to the manufacturers instructions. DNA was removed using the DNA-free DNA Removal Kit (Thermo Fisher Scientific, Massachusetts, USA) according to the manufacturers instructions and stored at 80C. The quality and quantity of total RNA was assessed by NanoDrop2000 spectrophotometer (Thermo Fisher Scientific, USA) and by RNA ScreenTape analyzer (average RIN for all samples was 8.2). The enriched RNA served as a template for constructions of cDNA libraries using random hexamers with qPCRBIO cDNA synthesis kit (PCR Biosystems, UK). RNA-seq libraries were prepared at the Crown Genomics institute of the Nancy and Stephen Grand Israel National Center for Personalized Medicine, Weizmann Institute of Science. Libraries were prepared using the INCPM-mRNA-seq protocol. Briefly, the polyA fraction (mRNA) was purified from 500ng of total input RNA followed by fragmentation and the generation of double-stranded cDNA. After Agencourt Ampure XP beads cleanup (Beckman Coulter), end repair, A base addition, adapter ligation and PCR amplification steps were performed. Libraries were quantified by Qubit (Thermo fisher scientific) and TapeStation (Agilent). Sequencing was done on a NovaSeq6000 instrument (Illumina) using an SP 200 cycles kit (paired read sequencing).