Identification and Characterization of Differentially Expressed microRNAs and Target Gene Related to Flax Stem Development

ABSTRACT

Flax fiber was one of the earliest natural fibers utilized by humans. Current research on flax fiber development is mainly focused on functions of key genes, with few reports investigating regulation of microRNA expression. Here, miRNA-seq was employed to explore miRNA expression profiles of middle stem tissues of high-fiber flax variety Agatha-MB and low-fiber flax variety White Flower during the rapid growth period (RGP) and mature green stage (MGS) to reveal miRNAs related to flax stem fiber development. The results revealed that 24 and 26 known miRNAs were identified that exhibited significant expression differences during MGSAgatha-MB-vs-RGPAgatha-MB and MGSWF-vs-RGPWF, respectively. Forty-one miRNA-target pairs exhibited inverse expression pattern changes as compared to our previously reported transcriptome profiling results. Pathway enrichment analysis of miRNA target genes showed that the plant hormone signal transduction pathway yielded the highest number of miRNA target genes in both varieties between the two developmental stages. To confirm the accuracy and reliability of these miRNA-seq results, qRT-PCR analysis of expression changes of six selected Lus-miRNA demonstrated similar trends as observed using miRNA-seq. These findings will enhance our understanding of miRNA regulatory mechanisms in flax stem development and will provide a foundation for future functional studies of these key miRNAs.

摘要

亚麻纤维是人类最早利用的天然纤维之一. 目前对亚麻纤维发育的研究主要集中在关键基因的功能上，很少有研究microRNA表达调控的报道. 利用miRNA-seq技术研究了高纤维亚麻品种Agatha-MB和低纤维亚麻品种白花在快速生长期（RGP）和成熟绿期（MGS）中茎组织miRNA的表达谱，揭示了与亚麻茎纤维发育相关的miRNA. 结果显示，在MGSAgatha-MB与RGPAgatha-MB以及MGSWF与RGPWF期间，分别鉴定出24个和26个已知的miRNAs表现出显著的表达差异. 与我们先前报道的转录组分析结果相比，41个miRNA靶对表现出反向表达模式变化. 对miRNA靶基因的途径富集分析表明，植物激素信号转导途径在两个发育阶段产生的miRNA靶基因在两个品种中最多. 为了证实这些miRNA-seq结果的准确性和可靠性，对6个选定Lus-miRNA表达变化的qRT-PCR分析显示了与使用miRNA-seq观察到的相似的趋势. 这些发现将增强我们对亚麻茎发育中miRNA调控机制的理解，并为这些关键miRNA的未来功能研究提供基础.

KEYWORDS:

• Flax
• microRNAs
• stem
• target gene
• fiber
• qRT-pcr

Keywords:

• 关键词
• 亚麻
• 微
• 茎
• 目的基因
• 纤维

Abbreviations

4CL: 4-coumarate-CoA ligase; ABI: Applied Biosystems; Agatha-MB: Agatha-Maintenance Breeding; BGI: Beijing Genomics Institute; CAD: cinnamyl alcohol dehydrogenase; CCoAOMT: caffeoyl CoA O-methyltransferase; CesA: cellulose synthase A; CNGBdb: China National GeneBank DataBase; COMT: catechol-o-methyl transferase; GAIP-B: gibberellic acid insensitive phloem B; GO: gene ontology; KEGG: kyoto encyclopedia of genes and genomes; lus-miR: linum usitatissimum microRNA; Lus-miRNA: linum usitatissimum microRNA; MGS: mature green stage; miRNA: microRNA; miRNA-seq: microRNA-seqencing; MYB: v-myb avian myeloblastosis viral oncogene homolog; PAGE: Polyacrylamide gel electrophoresis; PCR: polymerase chain reaction; piRNA: Piwi-interactiing RNA; qRT-PCR: quantitative RT-PCR; RGP: rapid growth period; RNA-seq: RNA-seqencing; siRNA: Small interfering RNA; snoRNA: small nucleolar RNA; sRNA: small RNA; UDP: uracil 5ʹ-diphosphate; UMI: unique molecular identifiers; WF: White Flower.

Additional information

Funding

This work was supported by the [National Natural Science Funds of China] under Grant [number 31801411](Ying Yu) and [Doctoral Research Fund of Guizhou University of Traditional Chinese Medicine] under Grant [number [2020]56](Ying Yu).