RNA-seq of FT3:cultured with 120 mM methanol and 30 mM sodium formate::Replicate 2
Total RNA (5g) extracted from Methylorubrum extorquens FT3 was used to construct the RNA-seq library. Ribosomal RNA was depleted using the Ribo-Zero Magnetic Kit, and RNA fragmentation was performed with the TruSeq Stranded RNA Sample Prep Kit. Double-stranded cDNA was then synthesized, incorporating dUTP in place of dTTP during second-strand synthesis. Indexed adapters were ligated to the double-stranded cDNA using the TruSeq Stranded RNA Sample Prep Kit. Subsequently, the cDNA second strand was degraded with Uracil-N-Glycosylase (UNG), and the library was enriched by 15 cycles of PCR amplification. Target fragments were size-selected using 2% Certified Low Range Ultra Agarose gel electrophoresis. Fragments were quantified via TBS380 (PicoGreen), and libraries were pooled based on concentration for sequencing. Cluster generation was performed via bridge PCR amplification on a cBot, followed by paired-end sequencing (2  150 bp) on the Illumina platform.