title: RNA Seq of Chryseobacterium sp. PMSZPI grown on hard agar.
design: Bacterial RNA Seq analysis was conducted to identify differentially expressed genes in hard agar against soft agar conditions in Chryseobacterium sp. PMSZPI. RNA was extracted from Chryseobacterium cell pellets and the integrity of the samples was assessed. RNA concentration was quantified and processed for ribodepletion. The rRNA-free and cleaned-up RNA was quantified on a Qubit fluorimeter. The enriched RNA was subjected to fragmentation, first-strand synthesis followed by second-strand synthesis. The double-stranded cDNA was ligated with an Illumina universal adapter and purified.  This ligated cDNA was subjected to indexing to enrich the adapter-ligated fragments. The final PCR product (sequencing library) was purified, followed by a library quality control check. Illumina-compatible sequencing libraries were quantified and its fragment size distribution was analyzed on . The libraries were paired-end sequenced on Illumina Novaseq 6000 sequencer (Illumina, San Diego, USA). Data Analysis Transcriptome analysis was performed by processing the raw data to remove low-quality data and adapter sequences. The high-quality reads were aligned against the reference genome to calculate expression. The expression data was further used for downstream analysis such as GO annotation, pathway assignments and differential gene expression using DESeq2.