Identification and characterization of microRNAs related to salt stress in broccoli, using high-throughput sequencing and bioinformatics analysis

Abstract

BackgroundMicroRNAs (miRNAs) are a new class of endogenous regulators of a broad range of physiological processes, which act by regulating gene expression post-transcriptionally. The brassica vegetable, broccoli (Brassica oleracea var. italica), is very popular with a wide range of consumers, but environmental stresses such as salinity are a problem worldwide in restricting its growth and yield. Little is known about the role of miRNAs in the response of broccoli to salt stress. In this study, broccoli subjected to salt stress and broccoli grown under control conditions were analyzed by high-throughput sequencing. Differential miRNA expression was confirmed by real-time reverse transcription polymerase chain reaction (RT-PCR). The prediction of miRNA targets was undertaken using the Kyoto Encyclopedia of Genes and Genomes (KEGG) Orthology (KO) database and Gene Ontology (GO)-enrichment analyses.ResultsTwo libraries of small (or short) RNAs (sRNAs) were constructed and sequenced by high-throughput Solexa sequencing. A total of 24,511,963 and 21,034,728 clean reads, representing 9,861,236 (40.23%) and 8,574,665 (40.76%) unique reads, were obtained for control and salt-stressed broccoli, respectively. Furthermore, 42 putative known and 39 putative candidate miRNAs that were differentially expressed between control and salt-stressed broccoli were revealed by their read counts and confirmed by the use of stem-loop real-time RT-PCR. Amongst these, the putative conserved miRNAs, miR393 and miR855, and two putative candidate miRNAs, miR3 and miR34, were the most strongly down-regulated when broccoli was salt-stressed, whereas the putative conserved miRNA, miR396a, and the putative candidate miRNA, miR37, were the most up-regulated. Finally, analysis of the predicted gene targets of miRNAs using the GO and KO databases indicated that a range of metabolic and other cellular functions known to be associated with salt stress were up-regulated in broccoli treated with salt.ConclusionA comprehensive study of broccoli miRNA in relation to salt stress has been performed. We report significant data on the miRNA profile of broccoli that will underpin further studies on stress responses in broccoli and related species. The differential regulation of miRNAs between control and salt-stressed broccoli indicates that miRNAs play an integral role in the regulation of responses to salt stress.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-014-0226-2) contains supplementary material, which is available to authorized users.

Identification of microRNAs and their targets in Finger millet by high throughput sequencing

“…miR159 and miR169 showed same expression patterns in root and shoot, studies have revealed that miR169 is responsive to abiotic stress such as drought, cold and salinity in different species (Ding et al, 2009;Li et al, 2013;Tian et al, 2014;Zhang et al, 2012). miR397 and miR398 also showed same expression levels in root and shoot.…”

MiRNA and phasiRNAs-mediated regulation of TIR-NBS-LRR defense genes inArabidopsis thaliana

“…Thus, in keeping with the designation of these miRNAs in other brassica species ( e.g. broccoli [46] ; A. lyrata miRBase Release 21 ), we will name these miRNAs as miR825-5p (22 nt, formerly miR825*) and miR825-3p (21 nt, formerly miR825)…”

“…MiR825-5p is conserved in Brassicaceae ( Fig. 1; [43, 46]). Using miR825-5p to search for potential target genes in the B. oleareacea genome in a recently created web browser (https://plantsmallrnagenes.science.psu.edu/ ; [77]), we found that several of its predicted targets, which display sequence similarity to Arabidopsis TNL genes, do accumulate significant amounts of 21 nt siRNAs, supporting the potential conservation of miR825-5p TNL regulatory network among Brassicaceae ( Fig.…”

Genome‐wide identification and characterization of Eutrema salsugineummicroRNAs for salt tolerance

“…Moreover, in our study, many miRNAs have a similar expression profile under salt stress in some species, but were regulated differently in other species. For example, miR160 levels were downregulated in the inflorescences of rice (Barrera-Figueroa et al 2012), B. juncea (Bhardwaj et al 2014), broccoli (Tian et al 2014) and S. europaea (Feng et al 2015) during salt stress, but upregulated in radish (Sun et al 2015) and H. caspica (Yang et al 2015). Similarly, miR390 was upregulated in Caragana intermedia (Zhu et al 2013), poplar (Populus tremula; Jia et al 2009), broccoli (Tian et al 2014), and radish (Sun et al 2015), but downregulated in B. juncea (Bhardwaj et al 2014).…”