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RNA sequencing, or RNA-Seq, is a high-throughput sequencing technology used to study the transcriptome of a cell. Transcriptomics helps to interpret the functional elements of a genome and identify the molecular constituents of an organism. Additionally, it also helps in understanding the development of an organism and the occurrence of diseases. 
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while...
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Multicellular, IVT-derived, unmodified human transcriptome for nanopore-direct RNA analysis.

Caroline A McCormick1, Stuart Akeson1, Sepideh Tavakoli1

  • 1Department of Bioengineering, Northeastern University, Boston, MA, 02115, USA.

Gigabyte (Hong Kong, China)
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Modification-free transcripts provide essential controls for Nanopore direct RNA sequencing (DRS). This study generated these controls from human cell lines, offering a valuable resource for RNA modification analysis.

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Area of Science:

  • Molecular Biology
  • Genomics
  • Bioinformatics

Background:

  • Nanopore direct RNA sequencing (DRS) allows for the detection of RNA modifications.
  • Modification-free transcripts are crucial controls for DRS, establishing a baseline for canonical nucleotides.
  • Generating these controls can be complex and introduce nanopore-specific analytical challenges.

Purpose of the Study:

  • To produce and characterize modification-free transcripts for Nanopore direct RNA sequencing (DRS).
  • To establish a versatile control resource for RNA modification analyses in human transcripts.
  • To investigate and interpret variations across different human cell lines.

Main Methods:

  • Produced direct RNA sequencing (DRS) datasets using modification-free transcripts.
  • Utilized *in vitro* transcription of cDNA derived from six immortalized human cell lines.
  • Characterized variations within and across cell lines for control data interpretation.

Main Results:

  • Generated high-quality DRS datasets from modification-free human transcripts.
  • Identified and characterized cell line-specific variations in the control data.
  • Demonstrated the utility of these datasets as a baseline for RNA modification studies.

Conclusions:

  • Modification-free transcripts generated via *in vitro* transcription serve as a practical control for Nanopore DRS.
  • The characterized datasets provide a valuable resource for the scientific community.
  • These controls aid in the accurate analysis of RNA modifications in human transcripts.