GSM4718793: YAK1 with Q578* variant replicate 4 [V4]; Saccharomyces cerevisiae; RNA-Seq
RNA extraction was done in two batches that each contained equal numbers of clones from the two groups (first batch: clones 1 and 2 of BY YAK1_wt and clones 1 and 2 of BY YAK1_Q578*, second batch: clones 3, 4 and 5 of BY YAK1_wt and clones 3, 4 and 5 of BY YAK1_Q578*). We used ZR Fungal/Bacterial RNA mini-prep kit with DNase I digestion (Zymo Research R2014), following the kit manual. RNA was eluted in 50 µl of RNase/DNase free water, quantified, and checked for integrity on an Agilent 2200 TapeStation. All RNA Integrity Numbers were higher than 9.5 and all concentrations were above 120 ng/µl. The RNA samples were stored at -20°C until use. Poly-A RNA selection was done using 550 ng of total RNA using NEBNext Poly(A) mRNA Magnetic Isolation Module (NEB E7490L), processing all samples in one batch. We prepared the library using NEB Ultra II Directional RNA library kit for Illumina (NEB E7760) used dual index primers (from NEBNext Multiplex Oligos for Illumina, NEB E7600S) for multiplexing, and amplified the library for ten cycles. The libraries were quantified using Qubit dsDNA HS Assay Kit (ThermoFisher Scientist Q32854) and pooled at equal mass for sequencing using 75-bp single-end reads on Illumina NextSeq 550 at UMGC.
GEO
GSM4718793