The full-length transcriptome of C. elegans using direct RNA sequencing.

Area of Science:

• Genomics
• Molecular Biology
• Developmental Biology

Background:

• Current transcriptome annotations often lack full-length isoform support due to short sequencing reads.
• Computational inference from short reads limits the accuracy of transcript isoform identification, particularly in complex organisms like Caenorhabditis elegans.

Purpose of the Study:

• To characterize the developmental polyadenylated transcriptome of Caenorhabditis elegans using long-read direct RNA sequencing.
• To identify novel splice isoforms and 3' untranslated regions (3' UTRs) with full-length transcript support.
• To analyze developmental variations in poly(A) tail lengths and their correlations with expression and 3' UTR length.

Main Methods:

• Application of nanopore-based direct RNA sequencing to C. elegans.
• Characterization of full-length mRNA transcripts, including splice isoforms and 3' UTRs.
• Analysis of poly(A) tail length dynamics across developmental stages.

Main Results:

• Identification of 23,865 splice isoforms across 14,611 genes, with 3,452 novel isoforms.
• Discovery of 16,342 3' untranslated region (UTR) isoforms, including 2,640 novel ones.
• Characterization of 28,858 full-length transcript isoforms and observed developmental variation in poly(A) tail lengths.

Conclusions:

• Long-read direct RNA sequencing provides unprecedented full-length transcriptome characterization, significantly improving gene annotation.
• The study reveals substantial novel transcript diversity, particularly in splice isoforms and 3' UTRs, in C. elegans.
• Developmental regulation of poly(A) tail length and its relationship with gene expression and UTR length are complex and dynamic.