Population-scale peach genome analyses unravel selection patterns and biochemical basis underlying fruit flavor

A narrow genetic basis in modern cultivars and strong linkage disequilibrium in peach (Prunus persica) has restricted resolution power for association studies in this model fruit species, thereby limiting our understanding of economically important quality traits including fruit flavor. Here, we present a high-quality genome assembly for a Chinese landrace, Longhua Shui Mi (LHSM), a representative of the Chinese Cling peaches that have been central in global peach genetic improvement. We also map the resequencing data for 564 peach accessions to this LHSM assembly at an average depth of 26.34× per accession. Population genomic analyses reveal a fascinating history of convergent selection for sweetness yet divergent selection for acidity in eastern vs. western modern cultivars. Molecular-genetics and biochemical analyses establish that PpALMT1 (aluminum-activated malate transporter 1) contributes to their difference of malate content and that increases fructose content accounts for the increased sweetness of modern peach fruits, as regulated by PpERDL16 (early response to dehydration 6-like 16). Our study illustrates the strong utility of the genomics resources for both basic and applied efforts to understand and exploit the genetic basis of fruit quality in peach.

Fine-tuning the performance of ddRAD-seq in the peach genome

The advance of Next Generation Sequencing (NGS) technologies allows high-throughput genotyping at a reasonable cost, although, in the case of peach, this technology has been scarcely developed. To date, only a standard Genotyping by Sequencing approach (GBS), based on a single restriction with ApeKI to reduce genome complexity, has been applied in peach. In this work, we assessed the performance of the double-digest RADseq approach (ddRADseq), by testing 6 double restrictions with the restriction profile generated with ApeKI. The enzyme pair PstI/MboI retained the highest number of loci in concordance with the in silico analysis. Under this condition, the analysis of a diverse germplasm collection (191 peach genotypes) yielded 200,759,000 paired-end (2 × 250 bp) reads that allowed the identification of 113,411 SNP, 13,661 InDel and 2133 SSR. We take advantage of a wide sample set to describe technical scope of the platform. The novel platform presented here represents a useful tool for genomic-based breeding for peach.

Genome-wide identification of HSF family in peach and functional analysis of PpHSF5 involvement in root and aerial organ development

Background Heat shock factors (HSFs) play important roles during normal plant growth and development and when plants respond to diverse stressors. Although most studies have focused on the involvement of HSFs in the response to abiotic stresses, especially in model plants, there is little research on their participation in plant growth and development or on the HSF (PpHSF) gene family in peach (Prunus persica). Methods DBD (PF00447), the HSF characteristic domain, was used to search the peach genome and identify PpHSFs. Phylogenetic, multiple alignment and motif analyses were conducted using MEGA 6.0, ClustalW and MEME, respectively. The function of PpHSF5 was confirmed by overexpression of PpHSF5 into Arabidopsis. Results Eighteen PpHSF genes were identified within the peach genome. The PpHSF genes were nonuniformly distributed on the peach chromosomes. Seventeen of the PpHSFs (94.4%) contained one or two introns, except PpHSF18, which contained three introns. The in silico-translated PpHSFs were classified into three classes (PpHSFA, PpHSFB and PpHSFC) based on multiple alignment, motif analysis and phylogenetic comparison with HSFs from Arabidopsis thaliana and Oryza sativa. Dispersed gene duplication (DSD at 67%) mainly contributed to HSF gene family expansion in peach. Promoter analysis showed that the most common cis-elements were the MYB (abiotic stress response), ABRE (ABA-responsive) and MYC (dehydration-responsive) elements. Transcript profiling of 18 PpHSFs showed that the expression trend of PpHSF5 was consistent with shoot length changes in the cultivar ‘Zhongyoutao 14’. Further analysis of the PpHSF5 was conducted in 5-year-old peach trees, Nicotiana benthamiana and Arabidopsis thaliana, respectively. Tissue-specific expression analysis showed that PpHSF5 was expressed predominantly in young vegetative organs (leaf and apex). Subcellular localization revealed that PpHSF5 was located in the nucleus in N. benthamiana cells. Two transgenic Arabidopsis lines were obtained that overexpressed PpHSF5. The root length and the number of lateral roots in the transgenic seedlings were significantly less than in WT seedlings and after cultivation for three weeks. The transgenic rosettes were smaller than those of the WT at 2–3 weeks. The two transgenic lines exhibited a dwarf phenotype three weeks after transplanting, although there was no significant difference in the number of internodes. Moreover, the PpHSF5-OE lines exhibited enhanced thermotolerance. These results indicated that PpHSF5 might be act as a suppresser of growth and development of root and aerial organs.

Functional Analysis of the Gibberellin 2-oxidase Gene Family in Peach

Peach (Prunus persica L. Batsch) trees grow vigorously and are subject to intense pruning during orchard cultivation. Reducing the levels of endogenous gibberellins (GAs) represents an effective method for controlling branch growth. Gibberellin 2-oxidases (GA2oxs) deactivate bioactive GAs, but little is known about the GA2ox gene family in peach. In this study, we identified seven PpGA2ox genes in the peach genome, which were clustered into three subgroups: C19-GA2ox-I, C19-GA2ox-II, and C20-GA2ox-I. Overexpressing representative genes from the three subgroups, PpGA2ox-1, PpGA2ox-5, and PpGA2ox-2, in tobacco resulted in dwarf plants with shorter stems and smaller leaves than the wild type. An analysis of the GA metabolic profiles of the transgenic plants showed that PpGA2ox-5 (a member of subgroup C19-GA2ox-II) is simultaneously active against both C19-GAs and C20-GAs,which implied that C19-GA2ox-II enzymes represent intermediates of C19-GA2oxs and C20-GA2oxs. Exogenous GA3 treatment of shoot tips activated the expression of all seven PpGA2ox genes, with different response times: the C19-GA2ox genes were transcriptionally activated more rapidly than the C20-GA2ox genes. GA metabolic profile analysis suggested that C20-GA2ox depletes GA levels more broadly than C19-GA2ox. These results suggest that the PpGA2ox gene family is responsible for fine-tuning endogenous GA levels in peach. Our findings provide a theoretical basis for appropriately controlling the vigorous growth of peach trees.

Different Roles of the Five Alcohol Acyltransferases in Peach Fruit Aroma Development

Peach (Prunus persica) fruit emit more than 100 volatile organic compounds. Among these volatiles, γ-decalactone is the key compound that contributes to peach aroma. The final step in lactones biosynthesis is catalyzed by alcohol acyltransferases (AATs). In this study, five AAT genes were isolated in the peach genome, and the ways that these genes contribute toward the peach aroma were studied. The sequence analysis of the five AATs showed PpAAT4 and PpAAT5 are truncated genes, missing important residues such as HXXXD. The expressions of PpAATs were investigated to identify the roles in creating the peach aroma. The results indicated that only PpAAT1 is highly expressed during γ-decalactone formation. A functional survey of the five PpAATs, using the oleaginous yeast expression system, suggested that only PpAAT1 significantly increased the γ-decalactone content, whereas the other four PpAATs did not significantly alter the γ-decalactone content. Enzyme assays on PpAATs heterologously expressed and purified from Escherichia coli indicated that only PpAAT1 could catalyze the formation of γ-decalactone. All results indicated that PpAAT1 is a more efficient enzyme than the other four PpAATs during the γ-decalactone biosynthesis process in peach fruit. The results from this study should help improve peach fruit aroma.

An integrated peach genome structural variation map uncovers genes associated with fruit traits

Background Genome structural variations (SVs) have been associated with key traits in a wide range of agronomically important species; however, SV profiles of peach and their functional impacts remain largely unexplored. Results Here, we present an integrated map of 202,273 SVs from 336 peach genomes. A substantial number of SVs have been selected during peach domestication and improvement, which together affect 2268 genes. Genome-wide association studies of 26 agronomic traits using these SVs identify a number of candidate causal variants. A 9-bp insertion in Prupe.4G186800, which encodes a NAC transcription factor, is shown to be associated with early fruit maturity, and a 487-bp deletion in the promoter of PpMYB10.1 is associated with flesh color around the stone. In addition, a 1.67 Mb inversion is highly associated with fruit shape, and a gene adjacent to the inversion breakpoint, PpOFP1, regulates flat shape formation. Conclusions The integrated peach SV map and the identified candidate genes and variants represent valuable resources for future genomic research and breeding in peach.

Identification of peach growth regulating-factors and their expression under UVB

Background: Growth-regulating factors (GRFs) are one of the most important plant-specific transcription factors with vital roles in multiple biological processes. GRFs have been identified in a variety of plant species, but a handful of research has addressed GRF genes in peach (Prunus persica).Results: Here, we report 46 members of the GRF family in four Rosaceae, divided into six subfamilies according to phylogeny, gene structure, and motif composition. We detected three collinear gene pairs generated from peach by whole-genome duplication or segmental duplication, but no tandem repeats were detected. Expression pattern analysis found that most PpGRFs were preferentially expressed in young tissues, At the sametime, multiple types cis-elements were observed in PpGRF promoters, and PpGRFs could positively respond to ultraviolet B-rays (UVB) and gibberellin (GA)treatments at the transcriptional level. Also, the content of GA3 and indole-3-acetic acid (IAA) changed significantly after UVB irradiation, indicating that GRFs might be involved in new shoot development in peach.Conclusions: This study identified 10 GRF genes in the peach genome and systematically analyzed their properties, thereby providing a foundation for researchers to have a better understanding of this gene family in peach. PpGRF 3, 4, 5, 6, 7, 9, and 10 positive responses to UVB and GA3 signals indicate that they can serve as candidate functional genes to further study how tree potential is regulated in peach.

Construction of an SNP-based high-density genetic map for Japanese plum in a Chinese population using specific length fragment sequencing

The Japanese plum (Prunus salicina Lindl.) is one of the most important stone fruit crops in China. High-density linkage map is valuable resources which enhance functional genomics and genetic breeding studies. So far several Japanese plum linkage maps have been reported using different kinds of molecular markers; however, the marker numbers and chromosome coverage are limited. Recently, a newly developed strategy which genome sequencing towards specific-locus amplified fragments (SLAF) markers, has been proven to be powerful for rapid genotyping of genome-wide markers and for high-density genetic map construction. In this study, SLAF was used to genotype markers with 114 F1 seedlings from the ‘09–16’ × ‘Fortune’ cross. Suitable SLAF markers (160,344 out of 343,436,902 pair-end reads) were chosen to conduct genetic map construction, 16.31% of which were polymorphic. The overall integrated map contained 3,341 high quality SLAFs and 720 loci that were grouped in eight genetic linkage groups with a total length of 869.9 cM and an average distance of 1.21 cM, and only five gaps with a genetic distance > 5 cM between adjacent markers occurred in linkage group (LG) 3 and LG6. The number of markers with each LG ranged from 82.3 cM (LG3) to 138.3 cM (LG1). Aligning the map against the peach reference genome sequence (Prunus persica L.) indicated a strictly co-linear relationship between the LGs and peach genome, demonstrating the markers on ours LGs were well ordered. Overall, our studies identified large-scale of genetic markers and constructed high-density linkage maps for Japanese plum, which will obviously provide a solid foundation for marker-assisted selection and sequence assembly of the Japanese plum reference genome.