title: GSM9282761: EAgNPs(ppk gene knockout) sample3; Escherichia coli str. K-12 substr. MG1655; RNA-Seq
library_construction_protocol: Total RNA was extracted from the selected biological samples using an RNA extraction kit (manufacturer details), following the manufacturer's protocol. RNA quantity and quality were assessed using a NanoDrop spectrophotometer (Thermo Fisher Scientific, USA) and an Agilent 2100 Bioanalyzer (Agilent Technologies, USA). The RNA integrity number (RIN) was verified to be above 8.0, ensuring suitability for downstream sequencing. RNA libraries were prepared using the Illumina TruSeq Stranded mRNA Library Prep Kit (Illumina, San Diego, CA), according to the manufacturer's instructions. Ribosomal RNA was depleted from the total RNA, followed by RNA fragmentation. First-strand cDNA synthesis was carried out using random hexamer primers. In the second-strand cDNA synthesis, dTTPs replaced dUTPs. After end repair, A-tailing, adapter ligation, size selection, amplification, and purification with Agencourt AMPure XP beads (Beckman Coulter, USA), the libraries were quantified using a Qubit fluorometer, and their quality was validated with the Agilent Bioanalyzer.