Comparative transcriptomic analysis reveals key components controlling spathe color in Anthurium andraeanum (Hort.)

Anthurium andraeanum (Hort.) is an important ornamental in the tropical cut-flower industry. However, there is currently insufficient information to establish a clear connection between the genetic model(s) proposed and the putative genes involved in the differentiation between colors. In this study, 18 cDNA libraries related to the spathe color and developmental stages of A. andraeanum were characterized by transcriptome sequencing (RNA-seq). For the de novo transcriptome, a total of 114,334,082 primary sequence reads were obtained from the Illumina sequencer and were assembled into 151,652 unigenes. Approximately 58,476 transcripts were generated and used for comparative transcriptome analysis between three cultivars that differ in spathe color (‘Sasha’ (white), ‘Honduras’ (red), and ‘Rapido’ (purple)). A large number of differentially expressed genes (8,324), potentially involved in multiple biological and metabolic pathways, were identified, including genes in the flavonoid and anthocyanin biosynthetic pathways. Our results showed that the chalcone isomerase (CHI) gene presented the strongest evidence for an association with differences in color and the highest correlation with other key genes (flavanone 3-hydroxylase (F3H), flavonoid 3’5’ hydroxylase (F3’5’H)/ flavonoid 3’-hydroxylase (F3’H), and leucoanthocyanidin dioxygenase (LDOX)) in the anthocyanin pathway. We also identified a differentially expressed cytochrome P450 gene in the late developmental stage of the purple spathe that appeared to determine the difference between the red- and purple-colored spathes. Furthermore, transcription factors related to putative MYB-domain protein that may control anthocyanin pathway were identified through a weighted gene co-expression network analysis (WGCNA). The results provided basic sequence information for future research on spathe color, which have important implications for this ornamental breeding strategies.

Comparative Transcriptome Analysis Revealed Two Alternative Splicing bHLHs Account for Flower Color Alteration in Chrysanthemum

‘Jimba’ is a white chrysanthemum cultivar, which occasionally and spontaneously produces red flower petals under natural cultivation due to cyanidin-based anthocyanin accumulation. To investigate the underlying mechanism of this process, a comparative transcriptome was analyzed between white and turning red ‘Jimba’. The structural and regulatory genes of anthocyanin pathway were significantly up-regulated in turning red ‘Jimba’. Among them, two alternative splicings, CmbHLH2 and CmbHLH2.1, showed the most significantly up-regulated in turning red tissue. Transiently over-expressed 35S::CmMYB6-CmbHLH2 strongly induced anthocyanin accumulation in ‘Jimba’ flower petals, while moderate amount of anthocyanin was detected when over-expressed 35S::CmMYB6-CmbHLH2.1. Both CmbHLH2 and CmbHLH2.1 could interact with CmMYB6 to activate CmDFR promoter according to Yeast two-hybrid and dual-luciferase assay. Moreover, CmMYB6-CmbHLH2 but not CmMYB6-CmbHLH2.1 could activate the CmbHLH2 promoter to provide positive feedback loop regulation. Taken together, it suggested that both CmbHLH2 and CmbHLH2.1 involved in regulation flower color alteration in turning red ‘Jimba’, and CmbHLH2 played a predominant role in this process.

Genome-Wide Analysis of MYB10 Transcription Factor in Fragaria and Identification of QTLs Associated with Fruit Color in Octoploid Strawberry

The genus Fragaria encompass fruits with diverse colors influenced by the distribution and accumulation of anthocyanin. Particularly, the fruit colors of strawberries with different ploidy levels are determined by expression and natural variations in the vital structural and regulatory genes involved in the anthocyanin pathway. Among the regulatory genes, MYB10 transcription factor is crucial for the expression of structural genes in the anthocyanin pathway. In the present study, we performed a genome wide investigation of MYB10 in the diploid and octoploid Fragaria species. Further, we identified seven quantitative trait loci (QTLs) associated with fruit color in octoploid strawberry. In addition, we predicted 20 candidate genes primarily influencing the fruit color based on the QTL results and transcriptome analysis of fruit skin and flesh tissues of light pink, red, and dark red strawberries. Moreover, the computational and transcriptome analysis of MYB10 in octoploid strawberry suggests that the difference in fruit colors could be predominantly influenced by the expression of MYB10 from the F. iinumae subgenome. The outcomes of the present endeavor will provide a platform for the understanding and tailoring of anthocyanin pathway in strawberry for the production of fruits with aesthetic colors.

Transcriptome Analysis Revealed the Mechanism by Which Exogenous ABA Increases Anthocyanins in Blueberry Fruit During Veraison

Blueberry (Vaccinium spp.) is a popular healthy fruit worldwide. The health value of blueberry is mainly because the fruit is rich in anthocyanins, which have a strong antioxidant capacity. However, because blueberry is a non-model plant, little is known about the structural and regulatory genes involved in anthocyanin synthesis in blueberries. Previous studies have found that spraying 1,000 mg/L abscisic acid at the late green stage of “Jersey” highbush blueberry fruits can increase the content of anthocyanins. In this experiment, the previous results were verified in “Brightwell” rabbiteye blueberry fruits. Based on the previous results, the anthocyanin accumulation process in blueberry can be divided into six stages from the late green stage to the mature stage, and the transcriptome was used to systematically analyze the blueberry anthocyanin synthesis process. Combined with data from previous studies on important transcription factors regulating anthocyanin synthesis in plants, phylogenetic trees were constructed to explore the key transcription factors during blueberry fruit ripening. The results showed that ABA increased the anthocyanin content of blueberry fruits during veraison. All structural genes and transcription factors (MYB, bHLH, and WD40) involved in the anthocyanin pathway were identified, and their spatiotemporal expression patterns were analyzed. The expression of CHS, CHI, DFR, and LDOX/ANS in ABA-treated fruits was higher in the last two stages of maturity, which was consistent with the change in the anthocyanin contents in fruits. In general, six MYB transcription factors, one bHLH transcription factor and four WD40 transcription factors were found to change significantly under treatment during fruit ripening. Among them, VcMYBA plays a major role in the regulation of anthocyanin synthesis in ABA signaling. This result preliminarily explained the mechanism by which ABA increases the anthocyanin content and improves the efficiency of the industrial use of blueberry anthocyanins.

Anthocyanin Genes Involved in the Flower Coloration Mechanisms of Cymbidium kanran

The Orchidaceae, otherwise known as orchids, is one of the largest plant families and is renowned for its spectacular flowers and ecological adaptations. Various polymorphisms of orchid flower colour can attract pollinators and be recognised as valuable horticultural ornamentals. As one of the longest historic cultured orchids, Cymbidium kanran has been domesticated for more than 2,500 years and is an ideal species to study coloration mechanisms because of plentiful variations in floral coloration and abundant traditional varieties. In this study, we used two distinct colour-type flowers of C. kanran as experimental materials to elucidate the mechanism of flower coloration. High-performance liquid chromatography (HPLC) analysis revealed that anthocyanins in purple-red-type flowers include three types of anthocyanidin aglycones, peonidin, malvidin, and cyanidin, whereas anthocyanins are lacking in white-type flowers. Through comparative transcriptome sequencing, 102 candidate genes were identified as putative homologues of colour-related genes. Based on comprehensive correlation analysis between colour-related compounds and gene expression profiles, four candidates from 102 captured genes showed a positive correlation with anthocyanidin biosynthesis. Furthermore, transient expression of CkCHS-1, CkDFR, and CkANS by particle bombardment confirmed that recovery of their expression completed the anthocyanin pathway and produced anthocyanin compounds in white-type flowers. Collectively, this study provided a comprehensive transcriptomic dataset for Cymbidium, which significantly facilitate our understanding of the molecular mechanisms of regulating floral pigment accumulation in orchids.

Molecular Evaluation of Kyoho Grape Leaf and Berry Characteristics Influenced by Different NPK Fertilizers

Fertilization, a fundamental aspect of a plant’s life, has been of great concern for agricultural specialists to minimize the yield gap between actual and potential yield. Around the globe, fertilizers with different NPK ratios are being used to attain a better yield of grape. To find the suitable commercially available fertilizer for quality grape production, a 2 years (2017–2018) study was conducted for the evaluation of 10 fertilizers with different NPK ratios. Commercial fertilizers included were Zhanlan (16:16:16), Garsoni (15:15:15), Acron (16:16:16), Norway (21:7:12), Peters 1 (30:10:10), Nutrivant (14:14:30), Peters 2 (20:20:20), UMAX (15:15:15), G2 (20:20:20), and Yara (15:15:15). The fertilizer application rate was 20 g plant−1, and each was applied at L-29, L-33, and L-36 phenological stages. Chlorophylls, carotenoids, macro/micronutrients in leaf, and anthocyanin derivatives in grape peel were evaluated. Expression levels of 24 genes, including nitrogen, phosphorous, potassium, and anthocyanin pathways in leaf, peel, and pulp were validated by qPCR at L-29, L-33, and L-36 stages. Results indicated that Norway (21:7:12) and Peters 1 (30:10:10) increased carotenoids, chlorophylls, and anthocyanins in leaves, while Zhanlan (16:16:16) improved fruit biochemical attributes, and anthocyanin (cyanidin, delphinidin, petunidin, malvidin, peonidin, and pelargonidin contents). However, a better grape yield was obtained by the application of Peters 1 (30:10:10). Potassium pathway genes were upregulated by Nutrivant (14:14:30), phosphorous pathway genes by Peters 2 (20:20:20), and nitrogen pathway genes by Peters 1 (30:10:10), while Nutrivant (14:14:30) upregulated anthocyanin pathway genes and simultaneously enhanced anthocyanin biosynthesis in berry peels. Results of two years’ study concluded that Peters 1 (30:10:10) was proved better to increase yield, while Zhanlan (14:14:30) was superior in improving anthocyanin biosynthesis.

The report of anthocyanins in the betalain-pigmented genus Hylocereus is not well evidenced and is not a strong basis to refute the mutual exclusion paradigm

Here we respond to the paper entitled “Contribution of anthocyanin pathways to fruit flesh coloration in pitayas” (Fan et al., BMC Plant Biol 20:361, 2020). In this paper Fan et al. 2020 propose that the anthocyanins can be detected in the betalain-pigmented genus Hylocereus, and suggest they are responsible for the colouration of the fruit flesh. We are open to the idea that, given the evolutionary maintenance of fully functional anthocyanin synthesis genes in betalain-pigmented species, anthocyanin pigmentation might co-occur with betalain pigments, as yet undetected, in some species. However, in absence of the LC-MS/MS spectra and co-elution/fragmentation of the authentic standard comparison, the findings of Fan et al. 2020 are not credible. Furthermore, our close examination of the paper, and re-analysis of datasets that have been made available, indicate numerous additional problems. Namely, the failure to detect betalains in an untargeted metabolite analysis, accumulation of reported anthocyanins that does not correlate with the colour of the fruit, absence of key anthocyanin synthesis genes from qPCR data, likely mis-identification of key anthocyanin genes, unreproducible patterns of correlated RNAseq data, lack of gene expression correlation with pigmentation accumulation, and putative transcription factors that are weak candidates for transcriptional up-regulation of the anthocyanin pathway.

Activation of PsMYB10.2 Transcription Causes Anthocyanin Accumulation in Flesh of the Red-Fleshed Mutant of ‘Sanyueli’ (Prunus salicina Lindl.)

Plum is one of the most important stone fruits in the world and anthocyanin-rich plums are increasingly popular due to their health-promoting potential. In this study, we investigated the mechanisms of anthocyanin accumulation in the flesh of the red-fleshed mutant of the yellow-fleshed plum ‘Sanyueli’. RNA-Seq and qRT-PCR showed that anthocyanin biosynthetic genes and the transcription factor PsMYB10.2 were upregulated in the flesh of the mutant. Functional testing in tobacco leaves indicated that PsMYB10.2 was an anthocyanin pathway activator and can activate the promoter of the anthocyanin biosynthetic genes PsUFGT and PsGST. The role of PsMYB10.2 in anthocyanin accumulation in the flesh of plum was further confirmed by virus-induced gene silencing. These results provide information for further elucidating the underlying mechanisms of anthocyanin accumulation in the flesh of plum and for the breeding of new red-fleshed plum cultivars.

A Multidisciplinary Fingerprinting Approach for Authenticity and Geographical Traceability of Portuguese Wines

The interest in developing reliable wine authenticity schemes is a hot-topic, especially for wines with recognized added-value. In order to accomplish this goal, two dimensions need to be considered: the grapevine variety determination and the geographical provenance. The aim of this study was to develop a multidisciplinary approach applicable to wines from the sub region Melgaço and Monção of the demarcated Vinho Verde region and from the demarcated Douro region. The proposed scheme consists on the use of DNA-based assays to detect Single Nucleotide Polymorphisms (SNPs) on three genes of the anthocyanin pathway (UFGT, F3H and LDOX) coupled with High-resolution melting (HRM) analysis aiming the varietal identification. The Alvarinho wines revealed to have the same haplotype using this marker set, demonstrating its applicability for genetic identification. In addition, to assess their geographical provenance, a multi-elemental approach using Sr and Pb isotopic ratios of wine, soil and bedrock samples was used. The isotopic data suggest a relation between Sr and Pb uptake by vine roots and soil’s texture and clay content, rather than with the whole rock’s isotopic ratios, but also highlights the potential of a discriminating method based on the combination of selected isotopic signatures.