BackgroundHydrogen cyanamide (HC) and pruning (P) have frequently been used to break dormancy in grapevine floral buds. However, the exact underlying mechanism remains elusive. This study aimed to address the early mode of action of these treatments on accumulation of reactive oxygen species (ROS) and reactive nitrogen species (RNS) and expression of related genes in the dormancy breaking buds of grapevine in the summer.ResultsThe budbreak rates induced by pruning (P), hydrogen cyanamide (HC), pruning plus hydrogen cyanamide (PHC) and water (control) after 8 days were 33, 53, 95, and 0 %, respectively. Clearly, HC was more effective in stimulating grapevine budbreak and P further enhanced its potency. In situ staining of longitudinal bud sections after 12 h of treatments detected high levels of ROS and nitric oxide (NO) accumulated in the buds treated with PHC, compared with HC or P alone. The amounts of ROS and NO accumulated were highly correlated with the rates of budbreak among these treatments, highlighting the importance of a rapid, transient accumulation of sublethal levels of ROS and RNS in dormancy breaking. Microarray analysis revealed specific alterations in gene expression in dormancy breaking buds induced by P, HC and PHC after 24 h of treatment. Relative to control, PHC altered the expression of the largest number of genes, while P affected the expression of the least number of genes. PHC also exerted a greater intensity in transcriptional activation of these genes. Gene ontology (GO) analysis suggests that alteration in expression of ROS related genes is the major factor responsible for budbreak. qRT-PCR analysis revealed the transient expression dynamics of 12 specific genes related to ROS generation and scavenge during the 48 h treatment with PHC.ConclusionOur results suggest that rapid accumulation of ROS and NO at early stage is important for dormancy release in grapevine in the summer, and the identification of the commonly expressed specific genes among the treatments allowed the construction of the signal transduction pathway related to ROS/RNS metabolism during dormancy release. The rapid accumulation of a sublethal level of ROS/RNS subsequently induces cell wall loosening and expansion for bud sprouting.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-016-0889-y) contains supplementary material, which is available to authorized users.

BackgroundDebaryomyces hansenii is one of the most salt tolerant species of yeast and has become a model organism for the study of tolerance mechanisms against salinity. The goal of this study was to identify key upregulated genes that are involved in its adaptation to high salinity.ResultsBy using forward subtractive hybridization we have cloned and sequenced DhAHP from D. hansenii that is significantly upregulated during salinity stress. DhAHP is orthologous to the alkly hydroperoxide reductase of the peroxiredoxin gene family, which catalyzes the reduction of peroxides at the expense of thiol compounds. The full-lengthed cDNA of DhAHP has 674 bp of nucleotide and contains a 516 bp open reading frame (ORF) encoding a deduced protein of 172 amino acid residues (18.3 kDa). D. hansenii Ahp is a cytosolic protein that belongs to the Ahp of the 1-Cys type peroxiredoxins. Phylogentically, the DhAhp and Candida albicans Ahp11 (Swiss-Prot: Q5AF44) share a common ancestry but show divergent evolution. Silence of its expression in D. hansenii by RNAi resulted in decreased tolerance to salt whereas overexpression of DhAHP in D. hansenii and the salt-sensitive yeasts Saccharomyces cereviasiae and Pichia methanolica conferred a higher tolerance with a reduced level of reactive oxygen species.ConclusionIn conclusion, for the first time our study has identified alkly hydroperoxide reductase as a key protein involved in the salt tolerance of the extremely halophilic D. hansenii. Apparently, this enzyme plays a multi-functional role in the yeast's adaptation to salinity; it serves as a peroxidase in scavenging reactive oxygen species, as a molecular chaperone in protecting essential proteins from denaturation, and as a redox sensor in regulating H2O2-mediated cell defense signaling.

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Background: Debaryomyces hansenii is a highly halotolerant yeast. The objective of this research is to investigate the genes that are involved in Debaryomyces hansenii for tolerance mechanisms against salinity. Methods: In this experiment, the gene DhSSA was cloned from D. hansenii and examined for its expression as induced by salt. The DhSSA gene is orthologous to the Saccharomyces cerevisiae SSA encoded HSP70 which cloned from D. hansenii induced by 2.5 M salt stress and significantly up-regulated during salinity stress. The full-length cDNA of DhSSA has 2,102 bp of nucleotide and contains a 1,926 bp Open Reading Frame (ORF) encoding a deduced protein of 642 amino acid residues (69.9 kDa). Results: Southern blot analysis indicated that DhSSA exists in the genome with one or two copy. Semi-quantitative RT-PCR and Real-time PCR results showed that expression of DhSSA is rapidly induced by salt and its expression increased with time reaching the highest level at 48 min before decline thereafter. Overexpression of DhSSA in Pichia methanolica conferred a higher stress tolerance. Immunocytological labeling reveals that DhSSA is a cytosolic protein. Conclusion: In summary, this study has cloned a salt-induced DhSSA gene from the halophilic yeast D. hansenii by subtraction hybridization. Functional analysis with overexpression transformants in P. methanolica suggest that the gene plays a protective role in the tolerance of D. hansenii to stresses.

The Cucurbit chlorotic yellows virus (CCYV) may infect many crops of Cucurbitaceae that leaves appear chlorosis or chlorotic spots and fruits yield low and are poor quality. Lately, the epidemic virus of CCYV has severely out-broken in field/greenhouse-grown cucurbits, and caused enormous losses of cucurbit industry. CCYV disease was first identified in Japan in 2004 and has rapidly spread in Asian, now CCYV has become to one of the most important epidemic virus of cucurbit crops. Until now there do not resistant cultivars released due to resistant gene of CCYV of cucurbits is not found. Recently resistant/tolerant plants of cucumber to CCYV have been identified by molecular diagnosis in my Lab. For identifying genes which involve CCYV resistance, we surveyed whole transcripts of cucumber (Cucumis sativus L.) inbred line 'Feng-tyan no.6', a tolerant line, through next generation sequencing (NGS). Total RNAs were extracted and whole transcriptome shotgun sequencing was performed using an Illumina platform. The gene expression level was calculated as FPKM. For differential expression analysis, CummeRbund was employed to perform statistical analyses of gene expression profiles. NGS detected a total of 19 differentially expressed genes (DEGs) with a higher abundance (>1-fold change with a p value <0.001) in the CCYV resistant transcriptome compared to that of the CCYV susceptible transcriptome. Among these DEGs relate to metabolic pathways, signalling pathways, DNA replication and cell cycle. The expression of these DEGs was further corroborated by PCR results. A comprehensive analysis of their gene expression profiles revealed that 3 genes (Gene ID: 101207198, 105435998 and 105436080) analysed were consistently resistance regulated by RT-PCR, indicating that the responses induced by CCYV. Moreover, the gene expression levels of 3 resistance regulated genes of 'Her torng no.11' of CCYV tolerant cultivars of cucumber was investigated at early and late stages of plants by real-time PCR, the results showed that the expression levels of 3 genes (Gene ID: 101207198, 105435998 and 105436080) of healthy plant are 400-826 folds, and are only 0.5-8.2 folds of disaster plants. The studies have provided the first glimpse of the molecular breeding of cucumber to resistant CCY disease.