Journal: Computational and Structural Biotechnology Journal

Article Title: Development of ONT-cappable-seq to unravel the transcriptional landscape of Pseudomonas phages

doi: 10.1016/j.csbj.2022.05.034

Figure Lengend Snippet: The ONT-cappable-seq technology. A. Schematic of the ONT-cappable-seq library preparation. After capping 5′ triphosporylated transcript ends with a desthiobiotin label, the primary transcripts are specifically enriched from the total RNA pool by streptavidin beads in the enriched library. The transcripts are polyA-tailed, converted to cDNA, barcoded and PCR amplified and loaded on the nanopore flow cell for sequencing. An unenriched control library is prepared in parallel. B . Bioinformatic pipeline implemented for LUZ7 phage transcripts analysis. The nanopore reads are base called and quality control is performed on the individual samples. Raw reads are trimmed, oriented and mapped to the reference genomes, followed by analysis of read distribution on genomic features and the elucidation of the transcriptional architecture of LUZ7. C . Comparison of Integrative Genomics Viewer (IGV) data tracks between the late transcriptomes of LUZ7 and LIT1 that were sequenced using ONT-cappable seq (top) or Illumina-based RNA-seq technology (bottom), respectively. Only the region of phage genomes that comprise two highly conserved genes in N4-like phage members encoding the major capsid protein (red) and N4 gp57-like protein (light blue), are shown in the IGV representations. While ONT-cappable-seq can accurately define TSSs (arrows) and TTSs (T) in LUZ7, genuine transcriptional boundaries are more challenging to identify from LIT1 classic RNA-seq data. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: C . Comparison of Integrative Genomics Viewer (IGV) data tracks between the late transcriptomes of LUZ7 and LIT1 that were sequenced using ONT-cappable seq (top) or Illumina-based RNA-seq technology (bottom), respectively.

Techniques: Amplification, Sequencing, Control, Comparison, RNA Sequencing

Journal: Computational and Structural Biotechnology Journal

Article Title: Development of ONT-cappable-seq to unravel the transcriptional landscape of Pseudomonas phages

doi: 10.1016/j.csbj.2022.05.034

Figure Lengend Snippet: Unusual intergenic transcription activity during the late infection stage of LUZ7 reveals putative small non-coding RNAs. Visualization in IGV of late infection transcripts mapping to the intergenic space between LUZ7 genes orf69 and orf70 reveals a condensed cluster of small transcripts, suggesting the presence of putative small non-coding RNAs. The most abundant RNA species are highlighted with blue bars and denominated as sRNA1, sRNA2, and sRNA3. sRNA candidates sRNA1 and sRNA2 share the same TSS (indicated with a green arrow) but use a different terminator sequence (indicated with a ‘T’). The sRNA3 lacks an annotated TSS and may arise due to processing events. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: C . Comparison of Integrative Genomics Viewer (IGV) data tracks between the late transcriptomes of LUZ7 and LIT1 that were sequenced using ONT-cappable seq (top) or Illumina-based RNA-seq technology (bottom), respectively.

Techniques: Activity Assay, Infection, Sequencing