Analysis of Transcriptome and Epitranscriptome in Plants Using PacBio Iso-Seq and Nanopore-Based Direct RNA...

Area of Science:

• Plant molecular biology and genomics.
• Transcriptomics and epitranscriptomics.

Background:

• Traditional short-read RNA sequencing (RNA-Seq) has limitations in analyzing full-length transcripts and RNA modifications.
• Long-read sequencing technologies offer a more comprehensive approach to transcriptome analysis.

Purpose of the Study:

• To summarize the applications of Nanopore and PacBio SMRT Iso-Seq in plant transcriptome analysis.
• To highlight the utility of direct RNA sequencing for studying RNA modifications and epitranscriptomes in plants.
• To discuss computational tools and limitations associated with these long-read methods.

Main Methods:

• Utilizing Oxford Nanopore Technologies (ONT) and Pacific BioSciences (PacBio) single-molecule real-time (SMRT) long-read sequencing.
• Direct RNA sequencing for capturing native RNA molecules and modifications.
• Bioinformatic analysis for identifying full-length isoforms, alternative splicing, fusion transcripts, and alternative polyadenylation.

Main Results:

• Long-read sequencing effectively identifies complex alternative splicing (AS), full-length splice variants, fusion transcripts, and alternative polyadenylation (APA) events in plants.
• Direct RNA sequencing provides insights into RNA modifications, advancing epitranscriptome research.
• These methods offer advantages over short-read approaches, requiring minimal sample processing.

Conclusions:

• Long-read sequencing is revolutionizing plant transcriptome analysis, enabling unprecedented insights into transcriptome complexity.
• These technologies are becoming the standard for plant transcriptomics, poised to uncover novel gene regulatory mechanisms.
• Future research will leverage these methods to understand plant development and stress responses.