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High-throughput gene-expression quantification of grapevine defense responses in the field using microfluidic dynamic arrays.

ABSTRACT: The fight against grapevine diseases due to biotrophic pathogens usually requires the massive use of chemical fungicides with harmful environmental effects. An alternative strategy could be the use of compounds able to stimulate plant immune responses which significantly limit the development of pathogens in laboratory conditions. However, the efficiency of this strategy in natura is still insufficient to be included in pest management programs. To understand and to improve the mode of action of plant defense stimulators in the field, it is essential to develop reliable tools that describe the resistance status of the plant upon treatment.We have developed a pioneering tool ("NeoViGen96" chip) based on a microfluidic dynamic array platform allowing the expression profiling of 85 defense-related grapevine genes in 90 cDNA preparations in a 4 h single run. Two defense inducers, benzothiadiazole (BTH) and fosetyl-aluminum (FOS), have been tested in natura using the "NeoViGen96" chip as well as their efficacy against downy mildew. BTH-induced grapevine resistance is accompanied by the induction of PR protein genes (PR1, PR2 and PR3), genes coding key enzymes in the phenylpropanoid pathway (PAL and STS), a GST gene coding an enzyme involved in the redox status and an ACC gene involved in the ethylene pathway. FOS, a phosphonate known to possess a toxic activity against pathogens and an inducing effect on defense genes provided a better grapevine protection than BTH. Its mode of action was probably strictly due to its fungicide effect at high concentrations because treatment did not induce significant change in the expression level of selected defense-related genes.The NeoViGen96" chip assesses the effectiveness of plant defense inducers on grapevine in vineyard with an excellent reproducibility. A single run with this system (4 h and 1,500 €), corresponds to 180 qPCR plates with conventional Q-PCR assays (Stragene system, 270 h and 9,000 €) thus a throughput 60-70 times higher and 6 times cheaper. Grapevine responses after BTH elicitation in the vineyard were similar to those obtained in laboratory conditions, whereas our results suggest that the protective effect of FOS against downy mildew in the vineyard was only due to its fungicide activity since no activity on plant defense genes was observed. This tool provides better understanding of how the grapevine replies to elicitation in its natural environment and how the elicitor potential can be used to reduce chemical fungicide inputs.

SUBMITTER: Dufour MC

PROVIDER: S-EPMC5120521 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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High-throughput gene-expression quantification of grapevine defense responses in the field using microfluidic dynamic arrays.

Dufour Marie-Cécile MC Magnin Noël N Dumas Bernard B Vergnes Sophie S Corio-Costet Marie-France MF

BMC genomics 20161122 1

<h4>Background</h4>The fight against grapevine diseases due to biotrophic pathogens usually requires the massive use of chemical fungicides with harmful environmental effects. An alternative strategy could be the use of compounds able to stimulate plant immune responses which significantly limit the development of pathogens in laboratory conditions. However, the efficiency of this strategy in natura is still insufficient to be included in pest management programs. To understand and to improve th ...[more]

PMID: 27875995

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Project description:Background: Only a subset of radically-resected pancreatic ductal adenocarcinoma (PDAC) patients benefit from chemotherapy, and identification of novel prognostic factors is warranted. Recently miRNAs emerged as diagnostic biomarkers and innovative therapeutic targets, while high-throughput arrays are opening new opportunities to evaluate whether they can also predict clinical outcome. The present study evaluated whether comprehensive miRNA expression profiling correlated with overall survival (OS) in resected PDAC patients. Methodology/Principal Findings: High-resolution miRNA profiles were obtained with the Toray’s 3D-GeneTM miRNA chip, detecting more than 1200 types of human miRNA. RNA was isolated from paraffin-embedded primary tumors of 26 radically resected stage-pT3N1 homogeneously treated patients (adjuvant gemcitabine 1000mg/m2/day, days-1/8/15, every 28days), carefully selected according to their outcome (OS<12 vs. OS>30 months, i.e. short/long-OS). Highly stringent statistics included t-test, distance matrix with Spearman-ranked correlation, and iterative approaches. Unsupervised hierarchical analysis revealed that PDAC specimens clustered according to their short/long-OS classification, while the feature selection algorithm RELIEF identified the top-4 discriminating miRNAs between the two groups. These miRNAs target more than 1500 transcripts, including 169 targeted by two or more of these miRNAs. MiR-211 emerged as the best discriminating miRNA, with significantly higher expression in long- vs. short-OS patients. The expression of this miRNA was subsequently assessed by quantitative-PCR in an independent cohort of laser microdissected tumors from 60 resected stage-pT3N1 PDAC patients treated with the same gemcitabine regimen. Patients with low miR-211 expression according to median value had a significantly shorter median OS (14.8, 95%CI=13.1-16.5, vs. 25.7 months, 95%CI=16.2-35.1, log-rank P=0.004). Multivariate analysis demonstrated that low miR-211 expression was an independent factor of poor prognosis (hazard ratio 2.3, P=0.03) after adjusting for all the factors influencing outcome. 19 samples, duplicated gene spots

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High-throughput gene-expression quantification of grapevine defense responses in the field using microfluidic dynamic arrays.

Publications

High-throughput gene-expression quantification of grapevine defense responses in the field using microfluidic dynamic arrays.

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