Abstract
The objective of the study is to reveal the freezing tolerance mechanisms of wheat by combining the emerging single-molecule real-time (SMRT) sequencing technology PacBio Sequel and Illumina sequencing. Commercial semiwinter wheat Zhoumai 18 was exposed to −6°C for 4 h at the four-leave stage. Leaves of the control group and freezing-treated group were used to perform cDNA library construction. PacBio SMRT sequencing yielded 51,570 high-quality isoforms from leaves of control sample of Zhoumai 18, encoded by 20,366 gene loci. In total, 73,695 transcript isoforms, corresponding to 23,039 genes, were identified from the freezing-treated leaves. Compared with transcripts from the International Wheat Genome Sequencing Consortium RefSeq v1.1, 57,667 novel isoforms were discovered, which were annotated 21,672 known gene loci, as well as 3,399 novel gene loci. Transcriptome characterization including alterative spliced events, alternative polydenylation sites, transcription factors, and fusion transcripts were also analyzed. Freezing-responsive genes and signals were uncovered and proved that the ICE-ERF-COR pathway and ABA signal transduction play a vital role in the freezing response of wheat. In this study, PacBio sequencing and Illumina sequencing were applied to investigate the freezing tolerance in common wheat, and the transcriptome results provide insights into the molecular regulation mechanisms under freezing treatment.
Highlights
Wheat (Triticum aestivum L.) is globally the third-largest food crop, next to maize and rice
Adapts and low-quality reads from the original sequencing data were filtered in SMRTlink v7.0; 30,066,483 (30.10 billion bases) and 31,782,257 (33.99 billion bases) subreads were generated from two samples, which formed 549,012 and 737,027 circular consensus sequences (CCSs), respectively (Table 1)
The full-length transcriptome is based on the PacBio Sequel three-generation sequencing platform, which can directly obtain complete transcripts containing 5, 3 UTR, and poly(A) tails without assembling short transcriptomic reads (Liu Q. et al, 2019)
Summary
Wheat (Triticum aestivum L.) is globally the third-largest food crop, next to maize and rice. It provides approximately 20% of human energy and is richer in protein and fat than other crops. Winter wheat can acquire cold tolerance through cold acclimation and vernalization, which usually takes ∼4 to 8 weeks, and during this period, plants can adapt to the gradually changing temperature. In recent years, warm winter and chilling in late spring occurred frequently; plants in key vegetative stages can be exposed to a very sudden subzero temperature when the winter-hardiness is almost lost (Li et al, 2015). RNA-seq has discovered several cold-related pathways and a series of key coldresponsive genes (Laudencia-Chingcuanco et al, 2011; Li et al, 2018; Pu et al, 2019)
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