T. thermophilius HB27c cells infected with Lalka27a phage; 5 min post infection
Total RNA was extracted using the RNA Purification from Lira Reagent Kit (Biolabmix, Russia). Libraries were constructed from 1 g of total RNA using the MGIEasy Fast RNA Library Prep Set (MGI Tech Co., Shenzhen, China) starting from RNA fragmentation (Section 3.2), without mRNA enrichment. First-strand cDNA was synthesized according to the manufacturers instructions; second-strand cDNA was generated with the Directional Second Strand Buffer (with dUTP) to retain strand specificity. End-repair/A-tailing was performed as recommended, followed by ligation of MGIEasy PF Adapters. Adapter-ligated DNA was size-selected and cleaned with the provided DNA Clean Beads, and libraries were amplified for 19 PCR cycles. Library quality and size distribution were assessed using the Agilent High Sensitivity D1000 ScreenTape Assay on a 4200 TapeStation System (Agilent Technologies, Santa Clara, CA, USA), and concentrations were determined with the QuDye ssDNA kit (Invitrogen, Thermo Fisher Scientific, USA). Barcoded libraries were pooled, converted to DNA nanoballs according to the manufacturers recommendations, and sequenced on a DNBSEQ-G400 in 2150 bp paired-end mode.