Characterization of the early gene expression profile inPopulus ussuriensisunder cold stress using PacBio SMRT sequencing integrated with RNA-seq reads

Abstract

Populus ussuriensis is an important and fast-growing afforestation plant species in north-eastern China. The whole-genome sequencing of P. ussuriensis has not been completed. Also, the transcriptional network of P. ussuriensis response to cold stress remains unknown. To unravel the early response of P. ussuriensis to chilling (3°C) stress and freezing (−3°C) stresses at the transcriptional level, we performed Single-Molecule Real-Time (SMRT) and Illumina RNA sequencing for P. ussuriensis. SMRT long-read isoform sequencing led to the identification of 29,243,277 subreads, and 575,481 circular consensus sequencing (CCS) reads. Approximately 50,910 high-quality (HQ) isoforms were generated, and 2,272 simple sequence repeats (SSRs), 8,086 long non-coding RNAs (lncRNAs) were identified. The Ca2+ content and abscisic acid (ABA) content in P. ussuriensis were significantly increased under cold stresses, while the value in freezing stress treatment group was significantly higher than chilling stress treatment group. A total of 49 genes that are involved in the signal transduction pathways related to perception and transmission of cold stress signals, such as Ca2+ signaling pathway, ABA signaling pathway, and MAPK signaling cascade, were found to be differentially expressed. In addition, 158 transcription factors (TFs) from 21 different families, such as MYB, WRKY, and AP2/ERF, were differentially expressed during chilling and freezing treatments. Moreover, the measurement of physiological indicators and bioinformatics observations demonstrated the altered expression pattern of genes involved in reactive oxygen species (ROS) balance, and sugar metabolism pathway during chilling and freezing stresses. This is the first time to report the early responses of P. ussuriensis to cold stress, which lays the foundation for future studies on the regulatory mechanisms in cold stress response. Besides, the full-length reference transcriptome of P. ussuriensis deciphered could be used in future studies on P. ussuriensis.

Characterization of the Gene Expression Profile Response to Drought Stress in Populus ussuriensis Using PacBio SMRT and Illumina Sequencing

“…These data were averaged from three biological replicates. In addition, according to the protocol outlined by Li et al [ 14 ], the free Ca 2+ content in root tip cells was measured using calcium indicator Flou-4 AM (Biyuntian, Shanghai, China), and observed through a confocal laser scanning microscope imager 700 (Carl Zeiss, Docuval, Germany).…”

“…Studies have exhaustively investigated molecular responses to drought stress in plants, such as Populus davidiana [ 8 ], Gossypium hirsutum and Gossypium tomentosum [ 9 ], Arabidopsis thaliana [ 10 ], and Oryza sativa [ 11 ]. TFs play an important role in the molecular response mechanism under drought stress, such as ABA-responsive element (ABRE), myeloblastosis (MYB), ethylene-responsive factor (ERF) and WRKY [ 12 , 13 , 14 ]. Additionally, synthesis of certain specific proteins, such as heat shock protein (HSP) and late embryogenesis abundant (LEA) protein, enhances the drought tolerance of plants [ 15 , 16 ].…”

ERF subfamily transcription factors and their function in plant responses to abiotic stresses

“…Cold stress, one of the most major abiotic stresses, can be generally categorized into chilling and freezing ( Li W. Y. et al., 2022 ). Both of them usually reduce the fluidity of cell membrane, affect the stability of proteins, break the intracellular ion homeostasis in plant ( Ding et al., 2019 ).…”

Physiological responses to drought stress of three pine species and comparative transcriptome analysis of Pinus yunnanensis var. pygmaea

“…Third-generation Pacific BioSciences (PacBio) single-molecule real-time (SMRT) sequencing does not need to interrupt RNA fragments, and direct reverse transcription can be used to obtain full-length cDNA [ 31 , 32 ]. Full-length transcripts that do not require assembly can be used to detect genes with AS events, APA events, lncRNAs and fusion transcripts [ 33 ]. Pacbio SMRT sequence is widely used to obtain long reads and assemble a high quality reference transcriptome.…”