title: GSM9211174: Kpn ST45, K3, R4; Klebsiella pneumoniae; RNA-Seq
library_name: GSM9211174
library_construction_protocol: RNA was extracted using the Macherey Nagel Trizol (ref:740971.250) according to the manufacturer's instructions and treated with DNase I provided in the kit. RNA concentration, quality, and integrity from four independent replicates were checked using the Invitrogen Qubit and the Agilent 2100 Bioanalyzer system. cDNA libraries were prepared from 1–3 µg of total RNA using the Illumina Total RNA Library Preparation Kit (Illumina, USA), following the manufacturer's protocol. To facilitate rRNA depletion, Ribo-Zero Plus Microbiome probes (Illumina) were used. Index barcodes were added by PCR for 13 cycles. Unbound adaptors and index primers were re-moved via purification with AMPure XP magnetic beads (Beckman Coulter, USA). The final libraries displayed an electrophoretic size distribution ranging from 250 to 900 bp, with a predominant peak at approximately 400 bp, as assessed on a 3500 Fragment Analyzer (Agilent Technologies, USA). Sequencing was performed on a NextSeq 2000 system using a P3 50-cycle flow cell (Illumina) to generate 67-nt single-end, dual-indexed reads.