A low-coverage 3' RNA-seq to detect homeolog expression in polyploid wheat

Area of Science:

• Genomics
• Transcriptomics
• Plant Science

Background:

• Distinguishing homeologous genes in allopolyploids is challenging for genomic analysis.
• Cost-efficient RNA sequencing (RNA-seq) is needed for large-scale studies in allopolyploids.

Purpose of the Study:

• To evaluate a low-cost 3' RNA-seq method (Lasy-Seq) for homeolog detection in allohexaploid bread wheat (Triticum aestivum).
• To assess the utility of 3' untranslated regions (UTRs) for differentiating homeologous genes.

Main Methods:

• Utilized 3' RNA-seq focusing on 3' UTRs with high mutation rates.
• Tested Lasy-Seq protocol on bread wheat.
• Compared HISAT2 mapping performance for 3' RNA-seq data.
• Extended 3' UTR annotation to improve homeolog detection.
• Validated findings with high-coverage conventional RNA-seq.

Main Results:

• HISAT2 demonstrated optimal performance for 3' RNA-seq, minimizing mapping errors and computational time.
• Extending 3' UTR annotation significantly enhanced homeolog detection.
• 3' RNA-seq successfully identified differentially expressed genes under cold stress, comparable to conventional RNA-seq.
• Downsampling revealed that even low sequencing depth (2 million reads) can detect over half of expressed homeologs.

Conclusions:

• Low-cost 3' RNA-seq is a viable tool for large-scale transcriptomic studies in allohexaploid wheat.
• This method shows potential for application in other allopolyploid species.
• 3' UTRs are valuable targets for homeolog identification in transcriptomic studies.