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Peach soft rot caused by Gilbertella persicaria is an economically important disease. Here, we report a high-quality complete and annotated genome sequence of G. persicaria strain TFLB-J, isolated from peach fruit in Yuanyang county of Henan Province, China. The assembly consists of 91 scaffolds with an estimated genome size of 33.59 Mb and N50 length of 0.92 Mb, encoding 13,296 predicted protein-coding genes. The whole-genome sequence could provide gene resources for further study of pathogenic effectors and comparative genomics of peach soft rot pathogens.

Peach (Prunus persica L.) is prone to chilling injury as exhibited by inhibition of the ethylene production, failure in softening, and the manifestation of internal browning. The basic leucine zipper (bZIP) transcription factors play an essential role in regulatory networks that control many processes associated with physiological, abiotic and biotic stress responses in fruits. Formerly, the underlying molecular and regulatory mechanism of (bZIP) transcription factors responsive to chilling injury in peach fruit is still elusive.In the current experiment, the solute peach 'Zhongyou Peach No. 13' was used as the test material and cold storage at low temperature (4 °C). It was found that long-term low-temperature storage induced the production of ethylene, the hardness of the pulp decreased, and the low temperature also induced ABA accumulation. The changes of ABA and ethylene in peach fruits during low-temperature storage were clarified. Since the bZIP transcription factor is involved in the regulation of downstream pathways of ABA signals, 47 peach bZIP transcription factor family genes were identified through bioinformatics analysis. Further based on RT-qPCR analysis, 18 PpbZIP genes were discovered to be expressed in refrigerated peach fruits. Among them, the expression of PpbZIP23 and PpbZIP25 was significantly reduced during the refrigeration process, the promoter analysis of these genes found that this region contains the MYC/MYB/ABRES binding element, but not the DRES/CBFS element, indicating that the expression may be regulated by the ABA-dependent cold induction pathway, thereby responding to chilling injury in peach fruit.Over investigation will provide new insights for further postharvest protocols related to molecular changes during cold storage and will prove a better cope for chilling injury.

Ethylene plays a critical role in peach (Prunus persica) fruit ripening; however, the molecular mechanism underlying ethylene-mediated aroma biosynthesis remains unclear. Here, we compared the difference in aroma-related volatiles and gene expression levels between melting-flesh (MF) and stony hard (SH) peach cultivars at S3, S4 I, S4 II, S4 III stages, and explored the relation between volatile biosynthesis related genes and ethylene response factor (ERF) genes. The concentration of fruity aromatic compounds such as lactones and terpenes increased significantly in MF peach during fruit ripening, while it was nearly undetectable in SH peach. LOX4 and FAD1 genes expressed concomitantly with ethylene emission and significantly downregulated by 1-MCP. Besides, 1-MCP treatment could sharply influence the fruity aromatic compounds, suggesting that these genes play key roles in volatile biosynthesis during fruit ripening. Furthermore, PpERF5 and PpERF7 could bind together to form a protein complex that enhanced the transcription of LOX4 more than each transcription factor individually. Overall, this work provides new insights into the transcriptional regulatory mechanisms associated with aroma formation during peach fruit ripening.

Background Ubiquitin ligases (E3) are the enzymes in the ubiquitin/26S proteasome pathway responsible for targeting proteins to the degradation pathway and play major roles in multiple biological activities. However, the E3 family and their functions are yet to be identified in the fruit of peach. Results In this study, genome-wide identification, classification and characterization of the E3 ligase genes within the genome of peach (Prunus persica) was carried out. In total, 765 E3 (PpE3) ligase genes were identified in the peach genome. The PpE3 ligase genes were divided into eight subfamilies according to the presence of known functional domains. The RBX subfamily was not detected in peach. The PpE3 ligase genes were not randomly distributed among the 8 chromosomes, with a greater concentration on the longer chromosomes. The primary mode of gene duplication of the PpE3 ligase genes was dispersed gene duplication (DSD). Four subgroups of the BTB subfamily never characterized before were newly identified in peach, namely BTBAND, BTBBL, BTBP and BTBAN. The expression patterns of the identified E3 ligase genes in two peach varieties that display different types of fruit softening (melting flesh, MF, and stony hard, SH) were analyzed at 4 different stages of ripening using Illumina technology. Among the 765 PpE3 ligase genes, 515 (67.3%) were expressed (FPKM > 1) in the fruit of either MF or SH during fruit ripening. In same-stage comparisons, 231 differentially expressed genes (DEGs) were identified between the two peach cultivars. The number of DEGs in each subfamily varied. Most DEGs were members of the BTB, F-box, U-box and RING subfamilies. PpE3 ligase genes predicted to be involved in ethylene, auxin, or ABA synthesis or signaling and DNA methylation were differentially regulated. Eight PpE3 ligase genes with possible roles in peach flesh texture and fruit ripening were discussed. Conclusions The results of this study provide useful information for further understanding the functional roles of the ubiquitin ligase genes in peach. The findings also provide the first clues that E3 ligase genes may function in the regulation of peach ripening.

Stony hard (SH) peach (Prunus persica L. Batsch) fruit does not release ethylene and has very firm and crisp flesh at ripening, both on- and off-tree. Long-term cold storage can induce ethylene production and a serious risk of chilling injury in SH peach fruit, however, the regulatory mechanism underlying ethylene production in stony hard peach is relatively unclear. In this study, we analyzed the phytohormone levels, fruit firmness, transcriptome, and lipidome changes in SH peach ‘Zhongtao 9’ (CP9) during cold storage (4 °C). The expression level of the ethylene biosynthesis gene PpACS1 and the content of ethylene in SH peach fruit were found to be upregulated during cold storage. A peak in ABA release was observed before the release of ethylene and the genes involved in ABA biosynthesis and degradation, such as zeaxanthin epoxidase (ZEP) and 8’-hydroxylase (CYP707A) genes, were specifically induced in response to low temperatures. Fruit firmness decreased fairly slowly during the first 20 d of refrigeration, followed by a sharp decline. Furthermore, the expression level of genes encoding cell wall metabolic enzymes, such as polygalacturonase, pectin methylesterase, expansin, galactosidase, and β-galactosidase, were upregulated only upon refrigeration, as correlated with the decrease in fruit firmness. Lipids belonging to 23 sub-classes underwent differential rearrangement during cold storage, especially ceramide (Cer), monoglycosylceramide (CerG1), phosphatidic acid (PA), and diacyglyceride (DG), which may eventually lead to ethylene production. Exogenous PC treatment provoked a higher rate of ethylene production. We suspected that the abnormal metabolism of ABA and cell membrane lipids promotes the production of ethylene under low temperature conditions, causing the fruit to soften. In addition, ERF transcription factors also play an important role in regulating lipid, hormone, and cell wall metabolism during long-term cold storage. Overall, the results of this study give us a deeper understanding of the molecular mechanism of ethylene biosynthesis during the postharvest storage of SH peach fruit under low-temperature conditions.

Members of the GOLVEN (GLV) gene family encode small secreted peptides involved in important plant developmental programs, including meristem maintenance and gravitropic responses. However, research on the functions of peptide hormones has mainly focused on various model plants, and little is known about their possible roles in peach fruit. Here, we identified eight PpGLV genes and analyzed their expression patterns in peach. PpGLV family genes share some similarities with Arabidopsis GLV family genes. PpGLV4 is responsive to 1-MCP and naphthalene acetic acid treatment. To investigate the role of PpGLV4 in detail, we treated peach callus with a synthetic PpGLV4 peptide. The upregulation of auxin and ethylene biosynthesis and signal transduction pathway genes in response to these treatment and the expression patterns of PpGLV family genes suggest that PpGLV4 may be involved in the crosstalk between auxin and ethylene signaling. The comprehensive data generated in this study improve our understanding of the roles of PpGLV peptides in peach.

The plant hormone ethylene regulates ripening in climacteric fruits. The phytohormone abscisic acid (ABA) affects ethylene biosynthesis, but whether ethylene influences ABA biosynthesis is unknown. To explore this possibility, we investigated the interactions between the ABA biosynthesis genes PpNCED2/3 and the ethylene response transcription factor PpERF3 in peach fruit. The ABA content increased during fruit maturation and reached a peak at stage S4 III. The increase was greatly inhibited by the ethylene inhibitor 1-MCP, which also suppressed PpERF3 expression. PpERF3 shared a similar expression profile with PpNCED2/3, encoding a rate-limiting enzyme involved in ABA biosynthesis, during fruit ripening. A yeast one-hybrid assay suggested that the nuclear-localized PpERF3 might bind to the promoters of PpNCED2/3. PpERF3 increased the expression of PpNCED2/3 as shown by dual-luciferase reporters, promoter-GUS assays and transient expression analyses in peach fruit. Collectively, these results suggest that ethylene promotes ABA biosynthesis through PpERF3’s regulation of the expression of ABA biosynthesis genes PpNCED2/3., Fruit ripening: Hormones go hand in hand Two hormones that regulate fruit ripening are more closely linked than previously thought, according to a study of ripening in peaches. Ethylene is a key ripening hormone in many fruits, and high ethylene levels turn on ethylene response factors (ERFs), genes that trigger production of sugars, pigments, and flavor compounds associated with ripening. Another hormone, abscisic acid (ABA), has recently been found to affect ripening, but its interaction with ethylene is unclear. Zhiqiang Wang and Guohuai Li at the Chinese Academy of Agricultural Sciences and coworkers investigated how ethylene and ABA interact during ripening. They found that as ethylene levels increased, ABA production was stimulated. Further investigation showed that ethylene directly triggered the ABA increase via ERFs. These results illuminate the fruit ripening process, and may help in finding ways to prolong fruit shelf life.

Background The green peach aphid (GPA), Myzus persicae (Sülzer), is a widespread phloem-feeding insect that significantly influences the yield and visual quality of peach [Prunus persica (L.) Batsch]. Single dominant gene (Rm3)-based resistance provides effective management of this invasive pest, although little is known about the molecular responses of plants to GPA feeding. Results To illustrate the molecular mechanisms of monogenic resistance in peach to young tissue-infecting GPAs, aphid-resistant/aphid-susceptible peach lines from a segregating population with Rm3/rm3 and rm3/rm3 genotypes were infested with GPAs for 3 to 72 h. Transcriptome analysis of the infested tissues identified 3854 differentially expressed genes (DEGs). Although the majority of the DEGs in the resistant line also responded to aphid attack in the susceptible line, the overall magnitude of change was greater in the resistant line than in the susceptible line. The enriched gene ontology of the 3854 DEGs involved in plant defence responses included redox situation, calcium-mediated signalling, transcription factor (e.g., WRKY, MYB, and ERF), MAPK signalling cascade, phytohormone signalling, pathogenesis-related protein, and secondary metabolite terms. Of the 53 genes annotated in a 460 kb interval of the rm3 locus, seven genes were differentially expressed between the aphid-resistant and aphid-susceptible peach lines following aphid infestation. Conclusions Together, these results suggest that the Rm3-dependent resistance relies mainly on the inducible expression of defence-related pathways and signalling elements within hours after the initiation of aphid feeding and that the production of specific secondary metabolites from phenylpropanoid/flavonoid pathways can have major effects on peach-aphid interactions. Electronic supplementary material The online version of this article (10.1186/s12864-018-5215-7) contains supplementary material, which is available to authorized users.

Flower and fruit colors are important agronomic traits. To date, there is no forward genetic evidence that the glutathione S-transferase (GST) gene is responsible for the white flower color in peach (Prunus persica). In this study, genetic analysis indicated that the white-flower trait is monogenetic, is recessive to the non-white allele, and shows pleiotropic effects with non-white-flowered types. The genetic locus underpinning this trait was mapped onto chromosome 3 between 0.421951 and 3.227115 Mb by using bulked segregant analysis in conjunction with whole-genome sequencing, and was further mapped between 0 and 1.178149 Mb by using the backcross 1 (BC1 ) population. Finally, the locus was fine-mapped within 535.974- and 552.027-kb intervals by using 151 F2 individuals and 75 individuals from a BC1 self-pollinated (BC1 S1 ) population, respectively. Pp3G013600, encoding a GST that is known to transport anthocyanin, was identified within the mapping interval. The analysis of genome sequence data showed Pp3G013600 in white flowers has a 2-bp insertion or a 5-bp deletion in the third exon. These variants likely render the GST non-functional because of early stop codons that reduce the protein length from 215 amino acids to 167 and 175 amino acids, respectively. Genetic markers based on these variants validated a complete correlation between the GST loss-of-function alleles and white flower in 128 peach accessions. This correlation was further confirmed by silencing of Pp3G013600 using virus-induced gene silencing technology, which reduced anthocyanin accumulation in peach fruit. The new knowledge from this study is useful for designing peach breeding programs to generate cultivars with white flower and fruit skin.

A massively parallel array of monoclonal antibodies enables rapid antibody development and target discovery across species., Antibodies are essential for elucidating gene function. However, affordable technology for proteome-scale antibody generation does not exist. To address this, we developed Proteome Epitope Tag Antibody Library (PETAL) and its array. PETAL consists of 62,208 monoclonal antibodies (mAbs) against 15,199 peptides from diverse proteomes. PETAL harbors binders for a great multitude of proteins in nature due to antibody multispecificity, an intrinsic antibody feature. Distinctive combinations of 10,000 to 20,000 mAbs were found to target specific proteomes by array screening. Phenotype-specific mAb-protein pairs were found for maize and zebrafish samples. Immunofluorescence and flow cytometry mAbs for membrane proteins and chromatin immunoprecipitation–sequencing mAbs for transcription factors were identified from respective proteome-binding PETAL mAbs. Differential screening of cell surface proteomes of tumor and normal tissues identified internalizing tumor antigens for antibody-drug conjugates. By finding high-affinity mAbs at a fraction of current time and cost, PETAL enables proteome-scale antibody generation and target discovery.

Peach (Prunus persica) is a typical climacteric fruit that produces ethylene rapidly during ripening, and its fruit softens quickly. Stony hard peach cultivars, however, do not produce large amounts of ethylene, and the fruit remains firm until fully ripe, thus differing from melting flesh peach cultivars. To identify the key proteins involved in peach fruit ripening, an antibody-based proteomic analysis was conducted. A mega-monoclonal antibody (mAb) library was generated and arrayed on a chip (mAbArray) at a high density, covering ~4950 different proteins of peach. Through the screening of peach fruit proteins with the mAbArray chip, differentially expressed proteins recognized by 1587 mAbs were identified, and 33 corresponding antigens were ultimately identified by immunoprecipitation and mass spectrometry. These proteins included not only important enzymes involved in ethylene biosynthesis, such as ACO1, SAHH, SAMS, and MetE, but also novel factors such as NUDT2. Furthermore, protein–protein interaction analysis identified a metabolon containing SAHH and MetE. By combining the antibody-based proteomic data with the transcriptomic and metabolic data, a mathematical model of ethylene biosynthesis in peach was constructed. Simulation results showed that MetE is an important regulator during peach ripening, partially through interaction with SAHH.

The auxin/indole acetic acid (Aux/IAA) genes encode a large family of short-lived proteins that regulate auxin signaling in plants. Previous attempts to understand the role of auxin during fruit development in peach (Prunus persica) found that expression of the Aux/IAA gene PpIAA19 was upregulated during fruit ripening, but the biological significance of the PpIAA19 protein is still unknown. The current study detected PpIAA19 transcripts in all peach plant tissues but found higher levels of accumulation in fruit S1 and S4II stages. During fruit ripening, the expression level of the PpIAA19 gene in melting flesh (MF) peach fruit was unsurprisingly higher than in stony hard (SH) peach fruit. Tomato (Solanum lycopersicum cv. ‘Micro-Tom’) was used as a model for investigating the phenotypic and molecular changes associated with over-expression of PpIAA19. This revealed that PpIAA19 was involved in regulating tomato lateral root number, stem elongation, parthenocarpy, and fruit shape. Molecular analyses of transgenic tomato plants over-expressing PpIAA19 indicated that the observed phenotypic changes were partially mediated by regulating the expression of auxin response genes. This result extends the functional characterization of PpIAA19 in tomato, and broadens our understanding of the role of auxin signaling in regulating plant growth and development.

The signaling pathways of both auxin and ethylene regulate peach fruit ripening via the Aux/IAA and ERF transcription factors, respectively. However, the molecular mechanisms that coordinate both auxin and ethylene signals during peach fruit ripening remain unclear. In this study, we show that PpIAA1 and PpERF4 act as key players in a positive feedback loop, and promote peach fruit ripening by directly binding to and enhancing the activity of target gene promoters. PpIAA1 increased the expression of the ethylene biosynthesis gene PpACS1. Furthermore, PpERF4 enhanced the transcription of PpACO1 and PpIAA1 genes by binding to their promoters. Additionally, PpIAA1 and PpERF4 bound to each other to form a complex, which then enhanced the transcription of abscisic acid biosynthesis genes (PpNCED2 and PpNCED3) and the fruit softening gene (PpPG1) to levels higher than those achieved by each transcription factor individually. Moreover, overexpression of PpIAA1 in tomato accelerated fruit ripening and shortened the fruit shelf-life by increasing the production of ethylene and the expression levels of ripening regulator genes. Collectively, these results advance our understanding of the molecular mechanisms underlying peach fruit ripening and softening via auxin and ethylene signaling pathways.

Ethylene is an essential regulator of fruit development. However, the role of this phytohormone in the development and maturation of peach (Prunus persica L. Batsch) fruit has not been systematically examined. In this study, through BLAST analysis of tomato (Solanum lycopersicum) protein sequences against the peach genome database and transcriptome analysis of all APETALA2/Ethylene Responsive Factor (AP2/ERF) genes in peach, we identified 15 ethylene signal transduction genes and 15 AP2/ERF genes and examined their expression levels in SH (stony hard) peaches and MF (melting flesh) peaches during fruit maturation. We investigated the expression patterns of six genes, including one ETR-like gene (Prupe.1G034300) and five AP2/ERF genes (Prupe.1G037700, Prupe.2G289500, Prupe.3G240000, Prupe.5G061800, and Prupe.7G194400), through quantitative RT-PCR expression analysis. The transcript levels of these genes were lower at the early stages of fruit development and increased dramatically at stage S4 in MF peaches, while they remained low in SH peaches, which mirrors the pattern of changes observed in ethylene production. Treatment of MF (‘Zhongyoutao 13’, ‘CN13’) fruit at stage S4 II with 1-MCP was ineffective in blocking ethylene production, and the expression of these genes was inhibited by 1-MCP treatment in ‘CN13’ on day 1, further demonstrating that these genes are closely related to fruit ripening. We detected a slight change in fruit firmness in ‘CN13’ after 1-MCP treatment, whereas ethylene production was not affected. By contrast, the transcript profile of gene Prupe.5G090800 exhibited the opposite trend. The comprehensive data generated in this study will improve our understanding of the ethylene receptor genes and ERF genes in peach and lay the foundation for further exploring the mechanisms underlying peach fruit ripening and softening.

The fruit skin pubescence of Prunus persica is an economically important characteristic and comprises the classification criteria. The mapping and identification of a complete linkage marker to the fruit skin trichome trait locus of peach fruit are critical for the molecular marker-assisted selection for peach/nectarine. In this study, the BC1 population was constructed from the parents “Zhongyou No. 4”, the recurrent parent, and “Baihuashanbitao”, the non-recurrent parent. Based on the 38 BC1 individuals’ phenotypes and their genotyping using next-generation sequencing, the G (Glabrous skin) locus of the gene was first identified between 14.099 and 16.721 Mb on chromosome 5. Using other individuals of this population, the gene was fine-mapped in the range of 481 kb with SNP markers. Based on the resequencing data of other cultivars (lines), the candidate SNP in the gene Prupe.5G196400 was obtained. Subsequently, the SNP marker was designed and applied to natural and hybrid peach populations. Via genotyping analysis, we confirmed co-segregation between the peach/nectarine phenotype, which was used in the identification of peach or nectarine with 100% accuracy.

Pectins are synthesized and secreted to the cell wall as highly methyl-esterified polymers and demethyl-esterified by pectin methylesterases (PMEs), which are regulated by pectin methylesterase inhibitors (PMEIs). PMEs and PMEIs are involved in pectin degradation during fruit softening; however, the roles of the PME and PMEI gene families during fruit softening remain unclear. Here, 71 PME and 30 PMEI genes were identified in the peach (Prunus persica) genome and shown to be unevenly distributed on all eight chromosomes. The 71 PME genes comprised 36 Type-1 PMEs and 35 Type-2 PMEs. Transcriptome analysis showed that 11 PME and 15 PMEI genes were expressed during fruit ripening in melting flesh (MF) and stony-hard (SH) peaches. Three PME and five PMEI genes were expressed at higher levels in MF than in SH fruit and exhibited softening-associated expression patterns. Upstream regulatory cis elements of these genes related to hormone response, especially naphthaleneacetic acid and ethylene, were investigated. One PME (Prupe.7G192800) and two PMEIs (Prupe.1G114500 and Prupe.2G279800), and their promoters were identified as potential targets for future studies on the biochemical metabolism and regulation of fruit ripening. The comprehensive data generated in this study will improve our understanding of the PME and PMEI gene families in peach. However, further detailed investigation is necessary to elucidate the biochemical function and regulation mechanism of the PME and PMEI genes during peach fruit ripening.

Cultivated peach (Prunus persica) is an important fruit species worldwide. The wild relatives in Prunus, such as P. mira, P. davidiana, P. kansuensis, P. ferganensis, and P. persica, are valuable for peach breeding, and early and accurate identification of parental and hybrid genotypes is critical. In this study, 20 representative accessions of peach germplasm from the National Germplasm Repository of Peach in China were used to select a set of 18 simple sequence repeat (SSR) markers for accurate species discrimination. Eight unknown peach samples were successfully identified using the SSR panel and species genotype database. Interspecific hybrid genotypes of P. persica × P. davidiana, P. persica × P. kansuensis, and P. persica × P. ferganensis were also analyzed reliably. The markers were amenable to high-throughput fluorescent labeling and capillary electrophoresis (CE) analysis, allowing rapid and efficient species identification. The practical method described in this study will facilitate peach breeding and germplasm management.

Antibodies are essential for elucidating the roles of genes decoded by genome sequencing. However, affordable technology for proteome-scale antibody generation does not exist. To address this, we developed the Proteome Epitope Tag Antibody Library (PETAL) and its array. PETAL consists of 62,208 mAbs against 15,199 peptides from diverse proteomes. PETAL harbors binders for a great multitude of proteins in nature due to antibody multispecificity, an intrinsic feature of an antibody. Distinctive combinations of 10,000-20,000 mAbs were found to target specific proteomes by array screening. Phenotype-specific mAb-target pairs were discovered for maize and zebrafish samples. Immunofluorescence and flow cytometry mAbs for human membrane proteins and ChIP-seq mAbs for transcription factors were identified from respective proteome-binding PETAL mAbs. Differential screening of cell surface proteomes of tumor and normal tissues discovered internalizing tumor antigens for antibody-drug conjugates. By discovering high affinity mAbs at a fraction of current time and cost, PETAL enables proteome-scale antibody generation and target discovery.

To characterize transcriptional regulation of the abscisic acid (ABA) signal transduction pathway during peach fruit early development and ripening, genes encoding the core components of the pathway, including 7 PpPYR (ABA receptor), 10 PpPP2C (type 2C protein phosphatase), and 7 PpSnRK2 (subfamily 2 of SNF1-related kinases), were identified from the peach (Prunus persica) reference genome. PpPYRs, PpPP2Cs, and PpSnRK2s from peach were putative homologous to proteins from tomato and Arabidopsis. Phylogenetic analysis showed that PpPYRs and PpSnRK2s were clustered into three subfamilies/subclasses. Transcript profile indicated that PpPYR2, PpPYR6, and PpPYR7 expression decreased sharply while ABA concentration increased during peach fruit ripening. Expression of PpPP2C4, PpPP2C9, and PpPP2C10 increased along with ABA concentration during fruit ripening. Expression of PpSnRK2.4 and PpSnRK2.6 also increased together with ABA concentration during fruit ripening. Furthermore, PpPP2C10 could interact with PpSnRK2.6 was determined by Yeast two-hybrid and BiFC analyses. The results will improve our understanding of ABA signal transduction genes in peach and lay the foundation for further exploring the mechanisms underlying peach fruit ripening.

Ethylene response factors (ERFs) are known to regulate fruit ripening. However, the ERF regulatory networks are not clear. In this study, we have shown that peach (Prunus persica) PpeERF2 regulates fruit ripening through suppressing the expression of two ABA biosynthesis genes (PpeNCED2, PpeNCED3) and a cell wall degradation gene (PpePG1). The transcript levels of PpeERF2 in fruit were opposite to that of PpeNCED2, PpeNCED3 and PpePG1 during ripening and in response to various ripening treatments. PpeERF2 was found to bind to the PpeNCED2, PpeNCED3 and PpePG1 promotors as demonstrated by yeast one-hybrid (Y1H) and EMSA assays; and further found to repress the promoter activities of the three genes in tobacco leaf tissues after Agrobacterium infiltration. Taken together, these results provide new information for a better understanding of the crosstalk network between ethylene signaling, cell wall degradation and ABA biosynthesis during fruit ripening.

In fleshy fruits, sugars (glucose, fructose, sucrose and sorbitol) are most important for fruit growth and development but also play a central role in fruit quality. In Peach (Prunus persica), we observed the sugars content at six developmental stages through high pressure liquid chromatography (HPLC). There was variation during fruit development and ripening, sucrose first increased then decreased; glucose and fructose first decreased, and then increased while sorbitol always decreased. In past, understanding about the mechanisms which control the sugar metabolism and accumulation in peach remains quite limited. We studied the transcript profiles of total 30 genes encoding key enzymes and transporters, which were involved in sucrose metabolism, resynthesis and transport. 13 putative genes including sucrose phosphatase synthase (PpSPS4 and PpSPS2), Hexokinases (PpHK1 and PpHK3), neutral/natural invertase (PpNINV1and PpNINV2), vacuolar acid invertase (PpVAINV2), sucrose transporter (PpSUT2), vacuolar glucose transporters (PpVGT1, PpVGT2 and PpVGT3) and tonoplast monosaccharides transporters (PpTMT1 and PpTMT2) highly expressed during peach fruit ripening. PpSPS4 had significantly positive correlation with glucose and fructose. Our results suggested that more sucrose accumulated in cytosol because of upregulation of (PpSPS4 and PpSPS2), that sucrose imported into vacuole through upregulation of PpSUT2, PpVGT1, PpVGT2, PpVGT3, PpTMT1 and PpTMT2 transporters, where cleaving of sucrose is done by upregulation of PpVAINV2. Hence, level of sucrose decreased as fruit ripe. As a result, the concentration of glucose and fructose increased at ripening stage (S4 II to S4 III). So, these marker genes can be involved in peach fruit ripening. This study will improve our understanding and lay foundation to explore the function of these genes and improvement of peach fruit quality.

Comparison of RNA-seq and qPCR gene expression data. (DOCX 444 kb)

Highlight PpYUC11 is a strong candidate gene for the control of the stony hard phenotype in peach., High concentrations of indole-3-acetic acid (IAA) are required for climacteric ethylene biosynthesis to cause fruit softening in melting flesh peaches at the late ripening stage. By contrast, the fruits of stony hard peach cultivars do not soften and produce little ethylene due to the low IAA concentrations. To investigate the regulation of IAA accumulation during peach ripening [the transition from stage S3 to stage S4 III (climacteric)], a digital gene expression (DGE) analysis was performed. The expression patterns of auxin-homeostasis-related genes were compared in fruits of the melting flesh peach ‘Goldhoney 3’ and the stony hard flesh peach ‘Yumyeong’ during the ripening stage. It is revealed here that a YUCCA flavin mono-oxygenase gene (PpYUC11, ppa008176m), a key gene in auxin biosynthesis, displayed an identical differential expression profile to the profiles of IAA accumulation and PpACS1 transcription: the mRNA transcripts increased at the late ripening stage in melting flesh peaches but were below the limit of detection in mature fruits of stony hard peaches. In addition, the strong association between intron TC microsatellite genotypes of PpYUC11 and the flesh texture (normal or stony hard) is described in 43 peach varieties, indicating that this locus may be responsible for the stony hard phenotype in peach. These findings support the hypothesis that PpYUC11 may play an essential role in auxin biosynthesis during peach fruit ripening and is a candidate gene for the control of the stony hard phenotype in peach.

Carotenoid content is an important determinant of pigmentation in citrus fruit. Phytoene synthase (PSY) is an important enzyme in the carotenoid synthesis pathway. In the present study, two similar but non-identical sequences transcripts (CsPsy1a and CsPsy1b) were isolated from sweet orange, and accumulated at a high level throughout the period of fruit maturation in both the peel and pulp. Although their transcript abundance was similar in the peel, Psy1a was the dominant form in the pulp. A functional analysis based on heterologous expression in Escherichia coli indicated that CsPsy1a was the more efficient converter of geranylgeranyl diphosphate to phytoene. In the pummelo cv. Gaophuang, which produces white fleshed fruit, Psy1b but not Psy1a transcript was detected. Expression of CsPsy1 genes and another seven carotenoid biosynthetic genes were studied in the pulp of ‘Anliu’ sweet orange and ‘Gaophuang’ pummelo, the variation in expression of genes associated with carotenoid synthesis could not explain the observed differences in carotenoid accumulation between orange and pummelo. Further site-directed mutagenesis in PSY1 suggested that maybe it was the number of AAT repeat units present in the gene which underlay variable activity of PSY1 in Citrus. These results have revealed the presence of diverse alleles of the Psy1 gene, with simple sequence repeats (SSRs) polymorphisms causing the change of enzymatic activity, are involved in the accumulation of total carotenoid in citrus fruits.

Oranges are an important nutritional source for human health and have immense economic value. Here we present a comprehensive analysis of the draft genome of sweet orange (Citrus sinensis). The assembled sequence covers 87.3% of the estimated orange genome, which is relatively compact, as 20% is composed of repetitive elements. We predicted 29,445 protein-coding genes, half of which are in the heterozygous state. With additional sequencing of two more citrus species and comparative analyses of seven citrus genomes, we present evidence to suggest that sweet orange originated from a backcross hybrid between pummelo and mandarin. Focused analysis on genes involved in vitamin C metabolism showed that GalUR, encoding the rate-limiting enzyme of the galacturonate pathway, is significantly upregulated in orange fruit, and the recent expansion of this gene family may provide a genomic basis. This draft genome represents a valuable resource for understanding and improving many important citrus traits in the future.

As a key component of tree architecture, plant height is an important agronomic trait in fruit trees. Reducing tree height is beneficial for increasing planting density, allowing higher yields, lowering costs and increasing the orchard’s lifespan. However, the genetic and molecular factors that regulate peach height are unknown. Here, we report a semi-dwarf peach mutant, which exhibits extremely shortened internodes at temperature below 30 °C. Genetic analysis indicated that this trait was controlled by a single dominant gene that we have named as temperature-sensitive semi-dwarf (Tssd). To map the Tssd locus, two DNA pools, each consisting of 50 mutant or wild-type siblings, were subjected to specific length amplified fragment sequencing. SLAF analysis followed by marker validation using Sanger sequencing and high-resolution melting genotyping located the Tssd gene in a region spanning approximately 750 kb between 2.35 and 3.10 Mb in scaffold 3 based on the reference peach genome sequence of ‘Lovell.’ Further SNPs were developed for fine mapping the locus, which spans an interval of 500 Kb and contains 69 predicted protein-coding gene models. The SNP markers flanking this interval can be applied in molecular identification of Tssd individuals. The result will provide information on how temperature regulates internode length in temperate fruit trees.

The phytoene synthase (PSY) gene product is involved in the carotenoid biosynthesis pathway. In this study, the promoter of the sweet orange gene CsPSY of sweet orange ‘Cara Cara’ (Citrus sinensis Osbeck [Cs]) was isolated and characterized. Its sequence included a number of regulatory elements predicted to be responsive to light and other environmental cues. The 926-bp region upstream of CsPSY was fused to the β-glucuronidase (GUS) reporter gene, and introduced as a transgene into Arabidopsis thaliana. The pattern of GUS expression in the transgenic plants showed that the CsPSY promoter drove expression in the young seedlings, the leaves, the roots and in parts of the flower, in a developmentally regulated fashion. A promoter deletion analysis revealed that the region −479 to −306 nt positively regulated expression, the region −306 to −93 nt was associated with negative regulation of expression and the region −46 to +21 nt maintained basal promoter activity. In the presence of sucrose, gene expression was regulated via a cis-acting promoter element(s). Two highly repetitive sequences (Sp1 and A-box), located in the region from −479 to −306 nt, may act as light-responsive elements regulating CsPSY transcription during photomorphogenesis.

Distinguishing a bud mutant from its parental genotype using conventional molecular markers is extremely difficult because they are thought to be genetically near-identical owing to the nature of somatic mutations that give rise to the bud mutant phenotype. In this study, array-comparative genome hybridization (array-CGH) was used to investigate the genetic variation between a sweet orange bud mutant “Hong Anliu” and its parental type “Anliu”. Surprisingly, 25 candidate genes were shown to exhibit gene dosage variations, with 18 being underrepresented and 7 overrepresented in the bud mutant in comparison with the parental type. These gene dosage variations were validated by quantitative PCR. Interestingly, however, the transcription levels of some of the corresponding genes were not significantly different between the two genotypes. The detection of gene dosage variations in at least 25 candidate genes between “Hong Anliu” and “Anliu” may explain in part the genetic difference responsible for the phenotypic deviation of the bud mutant from its parental cultivar. Our findings also suggest that array-CGH is a powerful technology that can be used for detection of genome-wide variations among near-identical genotypes such as bud mutants and their parental cultivars.

The nucleotide-binding site leucine-rich repeat (NBS-LRR) genes are the largest class of disease resistance genes in plants. However, our understanding of the evolution of NBS-LRR genes in Rutaceae fruit crops is rather limited. We report an evolutionary study of 103 NBS-encoding genes isolated from Poncirus trifoliata (trifoliate orange), Citrus reticulata (tangerine) and their F(1) progeny. In all, 58 of the sequences contained a continuous open reading frame. Phylogenetic analysis classified the 58 NBS genes into nine clades, eight of which were genus specific. This was taken to imply that most of the ancestors of these NBS genes evolved after the genus split. The motif pattern of the 58 NBS-encoding genes was consistent with their phylogenetic profile. An extended phylogenetic analysis, incorporating citrus NBS genes from the public database, classified 95 citrus NBS genes into six clades, half of which were genus specific. RFLP analysis showed that citrus NBS-encoding genes have been evolving rapidly, and that they are unstable when passed through an intergeneric cross. Of 32 NBS-encoding genes tracked by gene-specific PCR, 24 showed segregation distortion among a set of 94 F(1) individuals. This study provides new insight into the evolution of Rutaceae NBS genes and their behaviour following an intergeneric cross.

BACKGROUND: FJ72-1 navel orange and its bud mutant FJWC exhibit differences in melting texture character which are influenced mostly by cell wall metabolism. Here we compared the contents of water soluble pectin (WSP), protopectin, total pectin (TP), cellulose, and hemicellulose, activities of polygalaturonase (PG), pectin methylesterase (PME), pectate lyases (PL), cellulase (Cel) and gene expression levels of PG, PME, PL and Cel between the two cultivars. RESULTS: The content of cellulose and hemicellulose decreased progressively during fruit ripening. At the harvest time (230 DAF), the content of cellulose and hemicellulose in FJWC were obviously higher than those in FJ72-1; the WSP content, PG activities and its gene expression level in FJWC was lower than those in FJ72-1. Moreover, gene expression levels of PME and Cel in FJWC were only one-quarter of those in FJ72-1 at 230 DAF. CONCLUSION: The present work showed that the inferior melting character of FJWC attributed to the lower WSP, higher TP or protopectins, higher cellulose and hemicellulose than those in the pulp of FJ72-1 at harvest time. Lower expression levels of PG, PME and Cel at harvest time might be associated with the inferior melting character. Copyright © 2010 Society of Chemical Industry

A spontaneous sweet orange (Citrus sinenesis [L.] Osbeck) mutant 'Hong Anliu' is of high value due to lycopene accumulation in the pulp. In this study, we analyzed the proteomic alterations in the pulp of 'Hong Anliu' versus its wild type (WT) at four maturing stages by using 2-DE combined with MALDI-TOF-TOF MS. Among the 74 differentially expressed proteins identified, the majority are predicted to be involved in stress response, carbohydrate/energy metabolism and regulation, or protein fate, modification and degradation. Particularly, expression levels of six anti-oxidative enzymes were altered by the mutation; and assays of their respective enzymatic activities indicated an enhanced level of oxidative stress in 'Hong Anliu', implying a regulatory role of oxidative stress on carotenogenesis. This conclusion was further confirmed by our observation that treatment of fruit pulps with tert-butylhydroperoxide (a ROS progenitor) induced lycopene accumulation in 'Hong Anliu' only. Gene expression showed that genes predicted to function upstream of lycopene biosynthesis were generally upregulated in juice sacs, but downregulated in segment membranes in both 'Hong Anliu' and its WT. The result suggests an important role of post-transcriptional regulation on carotenogenesis since lycopene was induced in 'Hong Anliu' but not WT. The result also implies that carotenogenesis in juice sacs and segment membranes of citrus fruits may be regulated by different mechanisms.

Corona cage was an important research method to research corona effect. As design corona cage, the distance form cage wall to conductor, critical distance of AC charge, and corona cage length, and these questions should be resolved. Corona onset voltage, breakdown voltages were important coefficients to research conductor corona characteristics, and this was significant for transmission line design. To a certain diameter conductor, the diameter of corona cage should be small enough to make the conductor corona onset at a low voltage. Simultaneously, the gap between cage wall and conductor should be lager enough, so as to make breakdown voltage 50% higher than corona onset voltage. The critical distance of AC charge was calculated, and the length of corona cage was selected. The cross-section side width of the corona cage was 1.8m × 1.8m., and the measurement length of the cage was 3m, both of the protection length were 0.5m respectively. As in test, conductors were placed in at the center of the cage, and corona loss measurement system was adopted to measure conductors corona loss. Corona loss of conductor LGJ-630/45, LGJ-400/35, LGJ-300/40 were measured, and an excellent test effect was obtained.

An efficient method for analyzing seven sexual hormones (estradiol-17beta, estriol, estrone, testosterone, methyl-testosterone, progesterone and diethylstilbestrol) in essential oil by high performance liquid chromatography was developed. The samples were dissolved in n-hexane, then extracted with 90% methanol solution. The n-hexane layer was discarded and the methanol layer was cleaned-up twice with n-hexane. The target compounds were separated on an XTerra RP18 column (250 mm x 4.6 mm, 5 microm) using water-methanol-acetonitrile (50:30:20, v/v/v) as mobile phase in isocratic mode, and detected by a diode array detector (DAD) and a fluorescence detector (FLD). The flow rate was 1.0 mL/min. Estriol, estradiol-17beta, estrone, diethylstilbestrol were detected at 197 nm; progesterone, testosterone, methyl-testosterone were detected at 240 nm; estriol, estradiol-17beta, estrone were detected with the fluorescence detector simultaneously, the excitation and emission wavelengths were 280 nm and 310 nm, respectively. The average spiked recoveries for seven sexual hormones were above 93.0% except that of progesterone was 79.5%. The relative standard deviations of seven sexual hormones were from 0.90% to 1.89%. The linear ranges of the determination were from 0.010 mg/L to 1.0 mg/L. The method is simple and accurate for simultaneously analyzing the seven sexual hormones in essential oil.

A single-column method for the simultaneous determination of common cations and transition metals in real samples is proposed in this paper. Eleven cations (copper, lithium, sodium, ammonium, potassium, cobalt, nickel, magnesium, calcium, strontium and zinc) were separated and analyzed by means of ion chromatography using an isocratic elution with 2.5 mM methane sulfonic acid and 0.8 mM oxalic acid as mobile phase, IonPac SCS1 (250 mm x 4 mm I.D.) as the separation column and non-suppressed conductor detection. Optimized analytical conditions were further validated in terms of accuracy, precision and total uncertainty and the results showed the reliability of the IC method. The relative standard deviations (RSDs) of the retention time and peak area were less than 0.04 and 1.30%, respectively. The coefficients of determination for cations ranged from 0.9988 to 1.000. The method developed was successfully applied to determination of cations in samples of beer and bottled mineral water. The spiked recoveries for the cations were 94-106%. The method was applied to beer and beverage without interferences.