Late_BYIs rep 1
Total RNA was extracted from CGMMV-infected cucumber plants cv. KS. Leaves from two plants exhibiting either the early or the late post-recovery stage symptoms (two plants per each stage) and two un-inoculated control plants were subjected to dissections of uniform discs, 3mm in diameter, from either the newly identified bright yellow islands (see Results) or the corresponding dark surrounding tissues, in the symptomatic plants, and parallel dissections were performed on the control uninoculated plants. Total RNA was extracted from three combined discs, from each of the tested symptomatic features as well as the un-inoculated controls, using the GenEluteTM Mammalian Total RNA Miniprep Kit (Sigma-Aldrich) according to the manufacturer's instructions. Discs were ground in the lysis buffer using plastic sticks in 1.7ml Eppendorf vials. Total RNA was eluted into RNase free double-distilled water (DDW); DNA was removed using the DNA-freeTM DNA Removal Kit (Thermo Fisher Scientific, USA) according to the manufacturer's instructions and stored at -80 Celsius. The extracted RNA served for library constructions using the TruSeq RNA Library Prep (Illumina) commercial kit. RNA sequences containing poly-A tails were enriched using beads containing oligo-dT primers. The enriched RNA was used for constructions of cDNA libraries using random hexamers