Comprehensive transcriptome analysis of different potato cultivars provides insight into early blight disease caused by Alternaria solani.

BACKGROUND: Early blight, caused by the necrotrophic fungal pathogen Alternaria solani, is an economically important disease affecting the tuber yield worldwide. The disease is mainly controlled by chemical plant protection agents. However, over-using these chemicals can lead to the evolution of resistant A. solani strains and is environmentally hazardous. Identifying genetic disease resistance factors is crucial for the sustainable management of early blight but little effort has been diverted in this direction. Therefore, we carried out transcriptome sequencing of the A. solani interaction with different potato cultivars with varying levels of early blight resistance to identify key host genes and pathways in a cultivar-specific manner. RESULTS: In this study, we have captured transcriptomes from three different potato cultivars with varying susceptibility to A. solani,  namely Magnum Bonum, Désirée, and Kuras, at 18 and 36 h post-infection. We identified many differentially expressed genes (DEGs) between these cultivars, and the number of DEGs increased with susceptibility and infection time. There were 649 transcripts commonly expressed between the potato cultivars and time points, of which 627 and 22 were up- and down-regulated, respectively. Interestingly, overall the up-regulated DEGs were twice in number as compared to down-regulated ones in all the potato cultivars and time points, except Kuras at 36 h post-inoculation. In general, transcription factor families WRKY, ERF, bHLH, MYB, and C2H2 were highly enriched DEGs, of which a significant number were up-regulated. The majority of the key transcripts involved in the jasmonic acid and ethylene biosynthesis pathways were highly up-regulated. Many transcripts involved in the mevalonate (MVA) pathway, isoprenyl-PP, and terpene biosynthesis were also up-regulated across the potato cultivars and time points. Compared to Magnum Bonum and Désirée, multiple components of the photosynthesis machinery, starch biosynthesis and degradation pathway were down-regulated in the most susceptible potato cultivar, Kuras. CONCLUSIONS: Transcriptome sequencing identified many differentially expressed genes and pathways, thereby contributing to the improved understanding of the interaction between the potato host and A. solani. The transcription factors identified are attractive targets for genetic modification to improve potato resistance against early blight. The results provide important insights into the molecular events at the early stages of disease development, help to shorten the knowledge gap, and support potato breeding programs for improved early blight disease resistance.

Transcriptome profiling of two Moringa species and insights into their antihyperglycemic activity.

BACKGROUND: Moringa concanensis Nimmo (MC), a plant that resembles Moringa oleifera Lam. (MO), has less scientific information but has traditionally been used as a medicinal plant. Moringa species have long been known for their medicinal qualities, which include antioxidant, anti-inflammatory, anticancer, and antihyperglycemic effects. We investigated the antidiabetic potential of MC and MO species in this study by using transcriptome profiling, metabolite analysis, and in vitro assay studies. RESULTS: Our transcriptome analysis revealed the expression of enzymes involved in the biosynthesis of quercetin, chlorogenic acid, and benzylamine, all of which have previously been shown to have antidiabetic activity. We compared the expression patterns of five different tissues from MC and MO and it was found that the key enzymes involved in the biosynthesis of these compounds were highly expressed in leaf tissue. The expression estimated by MC transcriptome data in different tissues was verified using RT-qPCR analysis. The amount of these compounds was further quantified in the crude leaf extract of both species and found that MC had a higher abundance of quercetin and chlorogenic acid than MO. The crude leaf extract from both MC and MO were further tested in vitro, and the results demonstrated strong inhibitory activity for α-glucosidase and DPP-IV enzymes. Our findings suggest that compounds in leaf tissue, such as quercetin, benzylamine, and chlorogenic acid, could play a significant role in this antidiabetic activity. In addition, when comparing MO plants, we found that MC had a slightly higher effect in expression, abundance, and inhibitory activity. CONCLUSIONS: This study presents the first report of MC transcriptome data, as well as a comparison of its anti-diabetic activity to MO. Our analysis discussed the significance of leaf tissue in antidiabetic activity compared to other tissues of both species. Overall, this study not only provides transcriptome resources for Moringa species, but also sheds light on antidiabetic potential of both species.

The transcriptome enables the identification of candidate genes behind medicinal value of Drumstick tree (Moringa oleifera).

Moringa oleifera is a plant well-known for its nutrition value, drought resistance and medicinal properties. cDNA libraries from five different tissues (leaf, root, stem, seed and flower) of M. oleifera cultivar Bhagya were generated and sequenced. We developed a bioinformatics pipeline to assemble transcriptome, along with the previously published M. oleifera genome, to predict 17,148 gene models. Few candidate genes related to biosynthesis of secondary metabolites, vitamins and ion transporters were identified. Expressions were further confirmed by real-time quantitative PCR experiments for few promising leads. Quantitative estimation of metabolites, as well as elemental analysis, was also carried out to support our observations. Enzymes in the biosynthesis of vitamins and metabolites like quercetin and kaempferol are highly expressed in leaves, flowers and seeds. The expression of iron transporters and calcium storage proteins were observed in root and leaves. In general, leaves retain the highest amount of small molecules of interest.

Germination and stress tolerance of oats treated with pulsed electric field at different phases of seedling growth.

This study explores the impact of pulsed electric field (PEF) application on oat seedling growth and stress tolerance. PEF treatment (99 monopolar, rectangular pulses lasting 10 µs each, with a frequency of 13 Hz and a nominal electric field strength of 2250 V/cm) was applied at two growth stages: (i) when the seedlings had 0.2 cm roots emerging from the kernel, and (ii) when they had a 0.4 cm shoot emerging from the kernel. Post-treatment, the seedlings were hydroponically grown for 8 days. To induce stress, the hydroponic medium was augmented with PEG (15 %) to induce drought stress and NaCl (150 mM) to induce salinity stress. Results demonstrate that applying PEF improved the growth of the root and shoot of oat seedlings. This effect was more pronounced when applied to more developed seedlings. When PEF was applied during the later stage of germination, seedlings exposed to salinity stress showed enhanced shoot growth compared to the control. Under the studied conditions, the application of PEF had no impact on the growth of seedlings under drought stress.

Exploring the medicinally important secondary metabolites landscape through the lens of transcriptome data in fenugreek (Trigonella foenum graecum L.).

Fenugreek (Trigonella foenum-graecum L.) is a self-pollinated leguminous crop belonging to the Fabaceae family. It is a multipurpose crop used as herb, spice, vegetable and forage. It is a traditional medicinal plant in India attributed with several nutritional and medicinal properties including antidiabetic and anticancer. We have performed a combined transcriptome assembly from RNA sequencing data derived from leaf, stem and root tissues. Around 209,831 transcripts were deciphered from the assembly of 92% completeness and an N50 of 1382 bases. Whilst secondary metabolites of medicinal value, such as trigonelline, diosgenin, 4-hydroxyisoleucine and quercetin, are distributed in several tissues, we report transcripts that bear sequence signatures of enzymes involved in the biosynthesis of such metabolites and are highly expressed in leaves, stem and roots. One of the antidiabetic alkaloid, trigonelline and its biosynthesising enzyme, is highly abundant in leaves. These findings are of value to nutritional and the pharmaceutical industry.

Distinct Evolutionary Origins of Intron Retention Splicing Events in NHX1 Antiporter Transcripts Relate to Sequence Specific Distinctions in Oryza Species.

The genome of Asian cultivated rice (Oryza sativa L.) shows the presence of six organelle-specific and one plasma membrane (OsNHX1-7) NHX-type cation proton antiporters. Of these, vacuolar-localized OsNHX1 is extensively characterized. The genus Oryza consists of 27 species and 11 genome-types, with cultivated rice, diploid O. sativa, having an AA-type genome. Oryza NHX1 orthologous regions (gene organization, 5' upstream cis elements, amino acid residues/motifs) from closely related Oryza AA genomes cluster distinctly from NHX1 regions from more ancestral Oryza BB, FF and KKLL genomes. These sequence-specific distinctions also extend to two separate intron retention (IR) events involving Oryza NHX1 transcripts that occur at the 5' and 3' ends of the NHX1 transcripts. We demonstrate that the IR event involving the 5' UTR is present only in more recently evolved Oryza AA genomes while the IR event governing retention of the 13th intron of Oryza NHX1 (terminal intron) is more ancient in origin, also occurring in halophytic wild rice, Oryza coarctata (KKLL). We also report presence of a retro-copy of the OcNHX1 cDNA in the genome of O. coarctata (rOcNHX1). Preferential species and tissue specific up- or down-regulation of the correctly spliced NHX1 transcript/5' UTR/13th intron-retaining splice variants under salinity was observed. The implications of IR on NHX1 mRNA stability and ORF diversity in Oryza spp. is discussed.

A knowledge-driven protocol for prediction of proteins of interest with an emphasis on biosynthetic pathways.

This protocol describes a stepwise process to identify proteins of interest from a query proteome derived from NGS data. We implemented this protocol on Moringa oleifera transcriptome to identify proteins involved in secondary metabolite and vitamin biosynthesis and ion transport. This knowledge-driven protocol identifies proteins using an integrated approach involving sensitive sequence search and evolutionary relationships. We make use of functionally important residues (FIR) specific for the query protein family identified through its homologous sequences and literature. We screen protein hits based on the clustering with true homologues through phylogenetic tree reconstruction complemented with the FIR mapping. The protocol was validated for the protein hits through qRT-PCR and transcriptome quantification. Our protocol demonstrated a higher specificity as compared to other methods, particularly in distinguishing cross-family hits. This protocol was effective in transcriptome data analysis of M. oleifera as described in Pasha et al.•Knowledge-driven protocol to identify secondary metabolite synthesizing protein in a highly specific manner.•Use of functionally important residues for screening of true hits.•Beneficial for metabolite pathway reconstruction in any (species, metagenomics) NGS data.

Dataset for the combined transcriptome assembly of M. oleifera and functional annotation.

In this paper, we present the data acquired during transcriptome analysis of the plant Moringa oleifera [1] from five different tissues (root, stem, leaf, flower and seed) by RNA sequencing. A total of 271 million reads were assembled with an N50 of 2094 bp. The combined transcriptome was assessed for transcript abundance across five tissues. The protein coding genes identified from the transcripts were annotated and used for orthology analysis. Further, enzymes involved in the biosynthesis of select medicinally important secondary metabolites, vitamins and ion transporters were identified and their expression levels across tissues were examined. The data generated by RNA sequencing has been deposited to NCBI public repository under the accession number PRJNA394193 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA394193).

Identification of splice variant of OsGBF1 in Oryza sativa ssp. indica genotypes under salinity stress.

G-box-binding factors are plant transcription factors, involved in a wide range of biological processes including abiotic stress responses. In this study, we analyzed the expression of OsGBF1 during salt stress in two contrasting Oryza sativa spp. indica genotypes, Rasi and Tellahamsa. Two-day-old seedlings were exposed to NaCl stress under two different conditions. One set was exposed to 100 mM NaCl before transferring to 250 mM (induction stress), while another set was transferred directly to 250 mM (shock stress). During early induction stress, OsGBF1 was up-regulated in Rasi when compared to Tellahamsa. We cloned full-length OsGBF1 from these two genotypes, and analyzed the sequences. Our analysis indicated the presence of transcript variants, which are designated as OsGBF1a and 1b. OsGBF1b variant retained introns, which lead to the generation of premature termination codon. OsGBF1b transcript levels were not significantly different at 12-h of induction stress in Tellahamsa and Rasi. At 24-h of shock stress, OsGBF1b was up-regulated in both genotypes and the transcript was more in Rasi. Since, OsGBF1a and 1b are predicted to be splice variants, we examined expression pattern of OsSKIP, a splicing factor and component of the spliceosome. In induction stress, OsSKIP was up-regulated at 12- and 24-h in Rasi when compared to Tellahamsa. On the contrary, at 24-h shock stress, OsSKIP was down-regulated in Rasi when compared to Tellahamsa. It is possible that OsSKIP expression was increased in Rasi during induction stress for accurate splicing that could aid in tolerance. This is the first report on OsGBF1 splice variant and the variant could have specific functions linked to stress tolerance in rice.