Abstract
Alfalfa (Medicago sativa L.) is a high quality leguminous forage. Drought stress is one of the main factors that restrict the development of the alfalfa industry. High-throughput sequencing was used to analyze the microRNA (miRNA) profiles of alfalfa plants treated with CK (normal water), PEG (polyethylene glycol-6000; drought stress), and PEG + SNP (sodium nitroprusside; nitric oxide (NO) sprayed externally under drought stress). We identified 90 known miRNAs belonging to 46 families and predicted 177 new miRNAs. Real-time quantitative fluorescent PCR (qRT-PCR) was used to validate high-throughput expression analysis data. A total of 32 (14 known miRNAs and 18 new miRNAs) and 55 (24 known miRNAs and 31 new miRNAs) differentially expressed miRNAs were identified in PEG and PEG + SNP samples. This suggested that exogenous NO can induce more new miRNAs. The differentially expressed miRNA maturation sequences in the two treatment groups were targeted by 86 and 157 potential target genes, separately. The function of target genes was annotated by gene ontology (GO) enrichment and kyoto encyclopedia of genes and genomes (KEGG) analysis. The expression profiles of nine selected miRNAs and their target genes verified that their expression patterns were opposite. This study has documented that analysis of miRNA under PEG and PEG + SNP conditions provides important insights into the improvement of drought resistance of alfalfa by exogenous NO at the molecular level. This has important scientific value and practical significance for the improvement of plant drought resistance by exogenous NO.
Highlights
As a perennial forage, alfalfa has the advantages of good nutritional quality and high yield [1].It originated in Asia and Transcaucasia [2]
A total of 9 Small RNA (sRNA) libraries comprising three samples (CK: leaves with normal water, Polyethylene glycol-6000 (PEG): leaves with drought stress, PEG + SNP: leaves with drought stress and SNP treatment) were generated using the Illumina HiSeq 2500 platform, and the sequencing data of the three samples were averaged
The transport RNA (tRNA), miRNA, small nucleolar RNA (snoRNA), and ribosomal RNA (rRNA) reads were annotated as the clean sRNA in these libraries
Summary
As a perennial forage, alfalfa has the advantages of good nutritional quality and high yield [1]. It originated in Asia and Transcaucasia [2]. Alfalfa has been widely planted all over the world, and has been identified as a leading forage crop [3]. Alfalfa can improve the soil planting environment by fixing atmospheric nitrogen, reducing the concentration of diethylhexyl phthalate (DEHP) in the soil, thereby protecting the ecological environment [4]. The root system of alfalfa can effectively prevent water loss and soil erosion [5]. With the increasing demand for feed, alfalfa has been planted all over the country. As a global habitat problem, drought has seriously affected the yield and quality of alfalfa and restricted the development of the alfalfa industry [6]
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