BackgroundMost published genome sequences are drafts, and most are dominated by computational gene prediction. Draft genomes typically incorporate considerable sequence data that are not assigned to chromosomes, and predicted genes without quality confidence measures. The current Actinidia chinensis (kiwifruit) ‘Hongyang’ draft genome has 164 Mb of sequences unassigned to pseudo-chromosomes, and omissions have been identified in the gene models.ResultsA second genome of an A. chinensis (genotype Red5) was fully sequenced. This new sequence resulted in a 554.0 Mb assembly with all but 6 Mb assigned to pseudo-chromosomes. Pseudo-chromosomal comparisons showed a considerable number of translocation events have occurred following a whole genome duplication (WGD) event some consistent with centromeric Robertsonian-like translocations. RNA sequencing data from 12 tissues and ab initio analysis informed a genome-wide manual annotation, using the WebApollo tool. In total, 33,044 gene loci represented by 33,123 isoforms were identified, named and tagged for quality of evidential support. Of these 3114 (9.4%) were identical to a protein within ‘Hongyang’ The Kiwifruit Information Resource (KIR v2). Some proportion of the differences will be varietal polymorphisms. However, as most computationally predicted Red5 models required manual re-annotation this proportion is expected to be small. The quality of the new gene models was tested by fully sequencing 550 cloned ‘Hort16A’ cDNAs and comparing with the predicted protein models for Red5 and both the original ‘Hongyang’ assembly and the revised annotation from KIR v2. Only 48.9% and 63.5% of the cDNAs had a match with 90% identity or better to the original and revised ‘Hongyang’ annotation, respectively, compared with 90.9% to the Red5 models.ConclusionsOur study highlights the need to take a cautious approach to draft genomes and computationally predicted genes. Our use of the manual annotation tool WebApollo facilitated manual checking and correction of gene models enabling improvement of computational prediction. This utility was especially relevant for certain types of gene families such as the EXPANSIN like genes. Finally, this high quality gene set will supply the kiwifruit and general plant community with a new tool for genomics and other comparative analysis.Electronic supplementary materialThe online version of this article (10.1186/s12864-018-4656-3) contains supplementary material, which is available to authorized users.

Tomato, melon, grape, peach, and strawberry primarily accumulate soluble sugars during fruit development. In contrast, kiwifruit (Actinidia Lindl. spp.) and banana store a large amount of starch that is released as soluble sugars only after the fruit has reached maturity. By integrating metabolites measured by gas chromatography–mass spectrometry, enzyme activities measured by a robot-based platform, and transcript data sets during fruit development of Actinidia deliciosa genotypes contrasting in starch concentration and size, this study identified the metabolic changes occurring during kiwifruit development, including the metabolic hallmarks of starch accumulation and turnover. At cell division, a rise in glucose (Glc) concentration was associated with neutral invertase (NI) activity, and the decline of both Glc and NI activity defined the transition to the cell expansion and starch accumulation phase. The high transcript levels of β-amylase 9 (BAM9) during cell division, prior to net starch accumulation, and the correlation between sucrose phosphate synthase (SPS) activity and sucrose suggest the occurrence of sucrose cycling and starch turnover. ADP-Glc pyrophosphorylase (AGPase) is identified as a key enzyme for starch accumulation in kiwifruit berries, as high-starch genotypes had 2- to 5-fold higher AGPase activity, which was maintained over a longer period of time and was also associated with enhanced and extended transcription of the AGPase large subunit 4 (APL4). The data also revealed that SPS and galactinol might affect kiwifruit starch accumulation, and suggest that phloem unloading into kiwifruit is symplastic. These results are relevant to the genetic improvement of quality traits such as sweetness and sugar/acid balance in a range of fruit species.

This study identifies the developmental processes contributing to variation in green-fleshed kiwifruit (Actinidia deliciosa (A. Chev.) C.F. Liang et A.R. Ferguson var. deliciosa) fruit dry matter content (DM) and fresh weight (FW) by comparing genotypes with either high or low final DM. Results are compared with the model for fruit development, the tomato (Solanum lycopersicum L.). Differences in final composition were attributable to a higher rate of starch accumulation from 70 days after anthesis in high DM genotypes, with no other consistent differences in accumulation of soluble sugars or organic acids. High DM genotypes had 70% higher starch content and differed from low DM genotypes in the allocation of carbon between storage and other components. DM was negatively correlated with final fruit FW only in high DM genotypes, whereas starch was a constant proportion of dry weight (DW), suggesting a dilution effect rather than an interaction between fruit size and carbohydrate metabolism. Compared with tomato, the organic acids, particularly quinic acid, contributed more to estimated osmotic pressure during growth in FW than the soluble sugars, regardless of final composition or size. Seed mass per unit FW was highest in high DM genotypes, suggesting a previously unrecognised role for kiwifruit seeds in accumulation of carbohydrate by the pericarp. Anatomical comparisons also identified a role for differences in the packing of the two principal cell types, with an increased frequency of the larger cell type correlated with reduced DM. These genotypes demonstrate that kiwifruit differs from tomato in the role of starch as the principal stored carbohydrate, the reduced importance of dilution by growth in FW and the more minor role of the sugars compared with the organic acids during fruit development.

BackgroundPseudomonas syringae is a widespread bacterial species complex that includes a number of significant plant pathogens. Amongst these, P. syringae pv. actinidiae (Psa) initiated a worldwide pandemic in 2008 on cultivars of Actinidia chinensis var. chinensis. To gain information about the expression of genes involved in pathogenicity we have carried out transcriptome analysis of Psa during the early stages of kiwifruit infection.ResultsGene expression in Psa was investigated during the first five days after infection of kiwifruit plantlets, using RNA-seq. Principal component and heatmap analyses showed distinct phases of gene expression during the time course of infection. The first phase was an immediate transient peak of induction around three hours post inoculation (HPI) that included genes that code for a Type VI Secretion System and nutrient acquisition (particularly phosphate). This was followed by a significant commitment, between 3 and 24 HPI, to the induction of genes encoding the Type III Secretion System (T3SS) and Type III Secreted Effectors (T3SE). Expression of these genes collectively accounted for 6.3% of the bacterial transcriptome at this stage. There was considerable variation in the expression levels of individual T3SEs but all followed the same temporal expression pattern, with the exception of hopAS1, which peaked later in expression at 48 HPI. As infection progressed over the time course of five days, there was an increase in the expression of genes with roles in sugar, amino acid and sulfur transport and the production of alginate and colanic acid. These are both polymers that are major constituents of extracellular polysaccharide substances (EPS) and are involved in biofilm production. Reverse transcription-quantitative PCR (RT-qPCR) on an independent infection time course experiment showed that the expression profile of selected bacterial genes at each infection phase correlated well with the RNA-seq data.ConclusionsThe results from this study indicate that there is a complex remodeling of the transcriptome during the early stages of infection, with at least three distinct phases of coordinated gene expression. These include genes induced during the immediate contact with the host, those involved in the initiation of infection, and finally those responsible for nutrient acquisition.Electronic supplementary materialThe online version of this article (10.1186/s12864-018-5197-5) contains supplementary material, which is available to authorized users.

Kiwifruit (Actinidia spp.) is a recently domesticated fruit crop with several novel‐coloured cultivars being developed. Achieving uniform fruit flesh pigmentation in red genotypes is challenging. To investigate the cause of colour variation between fruits, we focused on a red‐fleshed Actinidia chinensis var. chinensis genotype. It was hypothesized that carbohydrate supply could be responsible for this variation. Early in fruit development, we imposed high or low (carbon starvation) carbohydrate supplies treatments; carbohydrate import or redistribution was controlled by applying a girdle at the shoot base. Carbon starvation affected fruit development as well as anthocyanin and carbohydrate metabolite concentrations, including the signalling molecule trehalose 6‐phosphate. RNA‐Seq analysis showed down‐regulation of both gene‐encoding enzymes in the anthocyanin and carbohydrate biosynthetic pathways. The catalytic trehalose 6‐phosphate synthase gene TPS1.1a was down‐regulated, whereas putative regulatory TPS7 and TPS11 were strongly up‐regulated. Unexpectedly, under carbon starvation MYB10, the anthocyanin pathway regulatory activator was slightly up‐regulated, whereas MYB27 was also up‐regulated and acts as a repressor. To link these two metabolic pathways, we propose a model where trehalose 6‐phosphate and the active repressor MYB27 are involved in sensing the carbon starvation status. This signals the plant to save resources and reduce the production of anthocyanin in fruits.

Exogenous application of a cytokinin-like compound forchlorfenuron (CPPU) can promote fruit growth, although often at the expense of dry matter (DM), an important indicator of fruit quality. Actinidia chinensis var. deliciosa ‘Hayward’ fruit are very responsive to CPPU treatments, but the mechanism underlying the significant fruit weight increase and associated decrease in DM is unclear. In this study, we hypothesised that CPPU-enhanced growth increases fruit carbohydrate demand, but limited carbohydrate supply resulted in decreased fruit DM. During fruit development, CPPU effects on physical parameters, metabolites, osmotic pressure and transcriptional changes were assessed under conditions of both standard and a high carbohydrate supply. We showed that CPPU increased fruit fresh weight but the dramatic DM decrease was not carbohydrate limited. Enhanced glucose and fructose concentrations contributed to an increase in soluble carbohydrate osmotic pressure, which was correlated with increased water accumulation in CPPU-treated fruit and up-regulation of water channel aquaporin gene PIP2.4 at 49 days after anthesis. Transcipt analysis suggested that the molecular mechanism contributing to increased glucose and fructose concentrations was altered by carbohydrate supply. At standard carbohydrate supply, the early glucose increase in CPPU fruit was associated with reduced starch synthesis and increased starch degradation. When carbohydrate supply was high, the early glucose increase in CPPU fruit was associated with a general decrease in starch synthesis but up-regulation of vacuolar invertase and fructokinase genes. We conclude that CPPU affected fruit expansion by increasing the osmotically-driven water uptake and its effect was not carbohydrate supply-limited.

The elevation of anthocyanin contents in fruits and vegetables is a breeding target for many crops. In some fruit, such as tomato, higher anthocyanin concentrations enhance storage and shelf life. In contrast, highly anthocyanic red-fleshed apples (Malus x domestica) have an increased incidence of internal browning flesh disorder (IBFD). To determine the mechanisms underlying this, ‘Royal Gala’ cultivar apples over-expressing the anthocyanin-related transcription factor (TF) MYB10 (35S:MYB10), which produces fruit with highly pigmented flesh, were compared with standard ‘Royal Gala’ Wild Type (WT) grown under the same conditions. We saw no incidence of IBFD in WT ‘Royal Gala’ but the over-expression of MYB10 in the same genetic background resulted in a high rate of IBDF. We assessed concentrations of potential substrates for IBDF and a comparison of metabolites in these apples showed that anthocyanins, chlorogenic acid, pro-cyanidins, flavon-3-ols, and quercetin were all higher in the MYB10 lines. For the flavol-3-ols sub-group, epicatechin rather than catechin was elevated in MYB10 lines compared with the control fruit. Internal ethylene concentrations were measured throughout fruit development and were significantly higher in 35S:MYB10 lines, and ethylene was detected at an earlier developmental stage pre-harvest. Expression analysis of key genes associated with ethylene biosynthesis (aminocyclopropane-1-carboxylic acid synthase and oxidase; ACS and ACO) and polyphenol oxidase (PPO) showed the potential for increased ethylene production and the mechanism for enhanced PPO-mediated browning. The expression of a transcription factor of the ethylene response factor (ERF) class, ERF106, was elevated in red flesh. Analysis of transcriptional activation by MYB10 showed that this transcription factor could activate the expression of apple ACS, ACO, and ERF106 genes. Our data show a link between the elevation of anthocyanin-related transcription factors and an undesirable fruit disorder. The accelerated advancement of maturity via premature ethylene induction has implications for the breeding and storage of these more highly pigmented plant products.

Genotypes were perceived differently from one another, and differences in fruit size and DM content were reflected in fruit sensorial properties. This study is unique in demonstrating interactions between fruit size, DM and sensory properties. These findings could be relevant not only to kiwifruit but to fruiting crop breeders in general, because of the demonstrated potential for effects of fruit size and DM content on sweetness, flavour and fruit texture.