Integrated Cytological, Physiological, and Transcriptome Analyses Provide Insight into the Albino Phenotype of Chinese Plum (Prunus salicina).

Albino seedlings that arise during seed reproduction can have a significant impact on plant growth and breeding. In this research, we present the first report of albino occurrences in the seed reproduction process of Prunus salicina and describe the cytological, physiological, and transcriptomic changes observed in albino seedlings. The albino seedlings which were observed in several plum cultivars exhibited abnormal chloroplast ultrastructure and perturbed stomatal structure. Compared to normal seedlings, the photosynthetic pigment contents in albino seedlings decreased by more than 90%, accompanied by significant reductions in several chlorophyll fluorescence parameters. Furthermore, substantially changed photosynthetic parameters indicated that the photosynthetic capacity and stomatal function were impaired in albino seedlings. Additionally, the activities of the antioxidant enzyme were drastically altered against the background of higher proline and lower ascorbic acid in leaves of albino seedlings. A total of 4048 differentially expressed genes (DEGs) were identified through transcriptomic sequencing, and the downregulated DEGs in albino seedlings were greatly enriched in the pathways for photosynthetic antenna proteins and flavonoid biosynthesis. GLK1 and Ftsz were identified as candidate genes responsible for the impaired chloroplast development and division in albino seedlings. Additionally, the substantial decline in the expression levels of examined photosystem-related chloroplast genes was validated in albino seedlings. Our findings shed light on the intricate physiological and molecular mechanisms driving albino plum seedling manifestation, which will contribute to improving the reproductive and breeding efforts of plums.

Transcriptome Profiles Reveal the Promoting Effects of Exogenous Melatonin on Fruit Softening of Chinese Plum.

In this study, we investigated the effect of exogenous melatonin (MT) on cell wall metabolism leading to Chinese plum (Prunus salicina Lindl.) fruit softening. Exogenous MT treatment increased the endogenous MT content in plum fruits before fruit ripening. However, in mature plum fruits, exogenous MT treatment decreased the fruit hardness, pulp hardness, fruit elasticity, contents of ion-bound pectin, covalently-bound pectin, hemicellulose, and cellulose, and activities of xyloglucan endotransglycosylase/hydrolase and endo-ß-1,4-glucanase, and increased the water-soluble pectin content, and activities of pectin methyl esterase, pectin lyase, polygalacturonase, ß-galactopyranosidase, and α-L-arabinofuranosidase. Transcriptome analysis revealed that the differentially expressed genes (DEGs) associated with cell wall metabolism in the exogenous MT-treated plum fruits were mainly enriched in the pentose and glucuronate interconversions, phenylpropanoid biosynthesis, cyanoamino acid metabolism, and galactose metabolism pathways. Analysis of these DEGs revealed that exogenous MT treatment affected the expression of genes regulating the cell wall metabolism. Overall, exogenous MT treatment promotes the fruit softening of Chinese plum.

Sugar composition and transcriptome analysis in developing 'Fengtang' plum (Prunus salicina Lindl.) reveal candidate genes regulating sugar accumulation.

Sweetness is an important attribute of fruit quality, which directly affects consumers' preference for fresh fruit and is mostly determined by carbohydrate composition. 'Fengtang' plum (Prunus salicina Lindl.) is recognized for its high soluble sugar content, but the sugar composition and the molecular mechanisms underlying sugar overproduction are not fully understood. In this work, the sugar components were analyzed using gas chromatography-mass spectrometry combined with transcription profiles from RNA-sequencing and Quantitative Real-time PCR during fruit development. The target metabolic group showed that sucrose was the dominant sugar component in mature fruit, followed by glucose, fructose, and sorbitol. Based on the transcriptome data and qRT-PCR validation, we identified 12 key structural genes that significantly responded to corresponding component accumulation: sucrose synthase (PsSUS4), sucrose phosphate synthase (PsSPS2), neutral invertase (PsNINV1/3/4), phosphoglucomutase (PsPGM1), UTP-glucose-1-phosphate uridylyl transferase (PsUGP1/2), hexose kinase (PsHXK1/3), sugar transport protein (PsSTP1), and Sugars Will Eventually be Exported Transporter (PsSWEET4). In which PsSUS4 and PsSPS2, whose encoding proteins immediately catalyze sucrose synthesis, were selected to be silenced using the virus-induced gene silencing technology. Silencing of PsSUS4 or PsSPS2 resulted in decreased sucrose content by 27.6% and 8%, respectively, compared with the control, verifying their important roles in sucrose accumulation. Subsequently, sugar metabolism networks in this high-sugar plum were constructed with 12 key structural genes, 72 putative transcription factors, and 4 major sugar components. These results might facilitate a better understanding of the molecular mechanisms of sugar accumulation in 'Fengtang' plum and provide a framework for future fruit quality improvement.

Development of PCR based SSR markers for Wilsonomyces carpophilus and a PCR based diagnosis protocol for the early detection of shot hole disease in stone fruit crops.

BACKGROUND: The conidial Ascomycota fungus Wilsonomyces carpophilus causing shot hole in stone fruits is a major constraint in the production of stone fruits worldwide. Shothole disease symptoms appear on leaves, fruits, and twigs. Successful isolation of the pathogen from different hosts on synthetic culture medium is a time consuming and tedious procedure for identification of the pathogen based on morpho-cultural characterization. METHODS AND RESULTS: The present research was carried out to develop a successful PCR based early detection protocol for the shot hole disease of stone fruits, viz., peach, plum, apricot, cherry, and almond using the pathogen specific SSR markers developed from the Wilsonomyces carpophilus genome using Genome-wide Microsatellite Analysing Tool package (GMATA) software. Diseased leaf samples of different stone fruits were collected from the SKUAST-K orchard and the pathogen was isolated on potato dextrose agar (PDA) medium and maintained on Asthana and Hawkers' medium with a total of 50 pathogen isolates comprised of 10 isolates each from peach, plum, apricot, cherry and almond. The DNA was extracted from both healthy and infected leaf samples of different stone fruits. The DNA was also extracted from the isolated pathogen cultures (50 isolates). Out of 2851 SSR markers developed, 30 SSRs were used for the successful amplification of DNA extracted from all the 50 pathogen isolates. These SSRs were used for the amplification DNA from shot hole infected leaf samples of different stone fruits, but the amplification was not observed in the control samples (DNA from healthy leaves), thus confirming the detection of this disease directly from the shot hole infected samples using PCR based SSR markers. To our knowledge, this forms the first report of SSR development for the Wilsonomyces carpophilus and their validation for the detection of shot hole disease directly from infected leaves. CONCLUSION: PCR based SSR makers were successfully developed and used for the detection of Wilsonomyces carpophilus causing shot hole disease in stone fruits including almond in nuts for the first time. These SSR markers could successfully detect the pathogen directly from the infected leaves of stone fruits namely peach, plum, apricot and cherry including almond from the nuts.

S-Locus Genotyping in Japanese Plum by High Throughput Sequencing Using a Synthetic S-Loci Reference Sequence.

Self-incompatibility in Prunus species is governed by a single locus consisting of two highly multi-allelic and tightly linked genes, one coding for an F-box protein-i.e., SFB in Prunus- controlling the pollen specificity and one coding for an S-RNase gene controlling the pistil specificity. Genotyping the allelic combination in a fruit tree species is an essential procedure both for cross-based breeding and for establishing pollination requirements. Gel-based PCR techniques using primer pairs designed from conserved regions and spanning polymorphic intronic regions are traditionally used for this task. However, with the great advance of massive sequencing techniques and the lowering of sequencing costs, new genotyping-by-sequencing procedures are emerging. The alignment of resequenced individuals to reference genomes, commonly used for polymorphism detection, yields little or no coverage in the S-locus region due to high polymorphism between different alleles within the same species, and cannot be used for this purpose. Using the available sequences of Japanese plum S-loci concatenated in a rosary-like structure as synthetic reference sequence, we describe a procedure to accurately genotype resequenced individuals that allowed the analysis of the S-genotype in 88 Japanese plum cultivars, 74 of them are reported for the first time. In addition to unraveling two new S-alleles from published reference genomes, we identified at least two S-alleles in 74 cultivars. According to their S-allele composition, they were assigned to 22 incompatibility groups, including nine new incompatibility groups reported here for the first time (XXVII-XXXV).

Transcriptome and metabolome analyses reveal anthocyanins pathways associated with fruit color changes in plum (Prunus salicina Lindl.).

Plum (Prunus salicina Lindl.) is one of the most widely cultivated and important fruit trees in temperate and cold regions. Fruit color is a significant trait relating to fruit quality in plum. However, its development mechanism has not been studied from the aspects of transcriptional regulation and metabolomic progress. To reveal the mechanism of fruit color developments in plums, we selected the fruits of two plum cultivars, 'Changli84' (Ch84, red fruit) and 'Dahuangganhe' (D, yellow fruit) as plant materials for transcriptome sequencing and metabolomic analysis were performed. Based on the data of transcriptome and metabolome at three fruit developmental stages, young fruit stage, color-change stage, and maturation stage, we identified 2,492 differentially expressed genes (DEGs) and 54 differential metabolites (DMs). The KEGG analysis indicated that "Flavonoid biosynthesis" was significantly enriched during three fruit development stages. Some DEGs in the "Flavonoid biosynthesis" pathway, had opposite trends between Ch84 and D, including chalcone synthase (CHS), dihydroflavonol 4-reductase (DFR) and flavonol synthase (FLS). Also, the genes encoding MYB-bHLH-WD (MBW) protein complexes, especially MYBs and bHLHs, showed a close relationship with plum fruit color. In the current study, DMs like procyanidin B1, cyanidin 3-glucoside, and cyanidin-3-O-alpha-arabinopyranoside were key pigments (or precursors), while the carotene and carotenoids did not show key relationships with fruit color. In conclusion, the anthocyanins dominate the color change of plum fruit. Carotenes and carotenoids might be related to the color of plum fruit, but do not play a dominate role.

Effects of Pollen Sources on Fruit Set and Fruit Characteristics of 'Fengtangli' Plum (Prunus salicina Lindl.) Based on Microscopic and Transcriptomic Analysis.

Adequate yield and fruit quality are required in commercial plum production. The pollen source has been shown to influence fruit set and fruit characteristics. In this study, 'Siyueli', 'Fenghuangli' and 'Yinhongli' were used as pollinizers of 'Fengtangli' plum. Additionally, self-pollination, mixed pollination, and open pollination were performed. We characterized the differences in pollen tube growth, fruit set and fruit quality among pollination combinations. 'Fengtangli' flowers pollinated by 'Fenghuangli' had more pistils with pollen tubes penetrating the ovary and the highest fruit set rate, while the lowest fruit set rate was obtained from self-pollination. In self-pollinated flowers, 33% of pistils had at least one pollen tube reaching the ovary, implying that 'Fengtangli' is partially self-compatible. Pollen sources affected 'Fengtangli' fruit size, weight, pulp thickness, soluble solids, and sugar content. Transcriptome analysis of 'Siyueli'-pollinated and 'Yinhongli'-pollinated fruits revealed 2762 and 1018 differentially expressed genes (DEGs) involved in the response to different pollen sources. DEGs were enriched in plant hormone signal transduction, starch and sucrose metabolism, and MAPK signaling pathways. Our findings provide a reference for the selection of suitable pollinizers for 'Fengtangli' plum and promote future research on the metaxenia effect at the molecular level.

Phylogenomic analysis and development of molecular markers for the determination of twelve plum cultivars (Prunus, Rosaceae).

BACKGROUND: Plums are one of the most important economic crops of the Rosaceae family and are produced all over the world. China has many local varieties, but the genomic information is limited for genetic studies. Here, we first sequenced, assembled, and analyzed the plastomes of twelve plum cultivars and developed molecular markers to distinguish them. RESULTS: The twelve plastomes of plum cultivars have a circular structure of 157,863-157,952 bp containing a large single-copy region (LSC) of 86,109-86,287 bp, a small copy region (SSC) of 18,927-19,031 bp, and two inverted repeats (IR) of 26,353-26,387 bp each. The plastomes of plum cultivars encode 131 genes, including 86 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. We detected 50, 54, 54, 53, 53, 50, 54, 54, 54, 49, 50, 54 SSRs in the twelve analyzed varieties, respectively. For repeat sequences, we identified 553 tandem repeats, 204 direct repeats, and 270 palindromic repeats. We also analyzed the expansion/contraction of IR regions. The genes rpl22, rps19, rpl2, ycf1, ndhF, and the trnH span on or near the boundary of IR and single-copy regions. Phylogenetic analysis showed that the twelve cultivars were clustered with the P. salicina and P. domestica. We developed eight markers LZ01 to LZ08 based on whole plastomes and nuclear genes and validated them successfully with six repetitions. CONCLUSIONS: The results obtained here could fill in the blanks of the plastomes of these twelve plum cultivars and provide a wider perspective based on the basis of the plastomes of Prunus to the molecular identification and phylogenetic construction accurately. The analysis from this study provides an important and valuable resource for studying the genetic basis for agronomic and adaptive differentiation of the Prunus species.

Whole Transcriptome Analyses of Apricots and Japanese Plum Fruits after 1-MCP (Ethylene-Inhibitor) and Ethrel (Ethylene-Precursor) Treatments Reveal New Insights into the Physiology of the Ripening Process.

The physiology of Prunus fruit ripening is a complex and not completely understood process. To improve this knowledge, postharvest behavior during the shelf-life period at the transcriptomic level has been studied using high-throughput sequencing analysis (RNA-Seq). Monitoring of fruits has been analyzed after different ethylene regulator treatments, including 1-MCP (ethylene-inhibitor) and Ethrel (ethylene-precursor) in two contrasting selected apricot (Prunus armeniaca L.) and Japanese plum (P. salicina L.) cultivars, 'Goldrich' and 'Santa Rosa'. KEEG and protein-protein interaction network analysis unveiled that the most significant metabolic pathways involved in the ripening process were photosynthesis and plant hormone signal transduction. In addition, previously discovered genes linked to fruit ripening, such as pectinesterase or auxin-responsive protein, have been confirmed as the main genes involved in this process. Genes encoding pectinesterase in the pentose and glucuronate interconversions pathway were the most overexpressed in both species, being upregulated by Ethrel. On the other hand, auxin-responsive protein IAA and aquaporin PIP were both upregulated by 1-MCP in 'Goldrich' and 'Santa Rosa', respectively. Results also showed the upregulation of chitinase and glutaredoxin 3 after Ethrel treatment in 'Goldrich' and 'Santa Rosa', respectively, while photosystem I subunit V psaG (photosynthesis) was upregulated after 1-MCP in both species. Furthermore, the overexpression of genes encoding GDP-L-galactose and ferredoxin in the ascorbate and aldarate metabolism and photosynthesis pathways caused by 1-MCP favored antioxidant activity and therefore slowed down the fruit senescence process.

Genetic Analysis of Prunus salicina L. by Random Amplified Polymorphic DNA (RAPD) and Intersimple Sequence Repeat (ISSR).

Background: Prunus salicina L. is an important fruit tree species of great economic value which is mainly distributed in the northern hemisphere. Methods: 25 samples of Prunus salicina L. were collected from 8 provinces in China, Japan, USA, and New Zealand. The genetic variations of these samples were characterized by the random amplified polymorphic DNA (RAPD) and intersimple sequence repeat (ISSR) technique, respectively, and in combination. Results: Totally, 257 RAPD bands ranging 200∼2300 bp was found, and 81.59% of these bands were polymorphic. ISSR analysis identified 179 bands ranging 300∼2500 bp, and 87.74% of the bands were polymorphic. ISSR results showed that the similarity coefficient index between samples P10 (Maihuangli in Anhui, Chin) and P13 (Longyuanqiuli in Heilongjiang, China) was lowest, while that between samples P10 (Maihuangli in Anhui, Chin) and P15 (Baili in Japan) was highest. Combined analysis of RAPD and ISSR demonstrated that the similarity coefficient index between samples P4 (Qiepili in Ningbo, Zhejiang, China) and P13 (Longyuanqiuli in Heilongjiang, China) was lowest, while that between samples P19 (Laroda in USA) and P20 (Red heart in USA) was highest. Conclusion: RAPD combined with ISSR analysis can be used for genetic characterization of Prunus L. species.

The genome of low-chill Chinese plum "Sanyueli" (Prunus salicina Lindl.) provides insights into the regulation of the chilling requirement of flower buds.

Chinese plum (Prunus salicina Lindl.) is a stone fruit that belongs to the Prunus genus and plays an important role in the global production of plum. In this study, we report the genome sequence of the Chinese plum "Sanyueli", which is known to have a low-chill requirement for flower bud break. The assembled genome size was 282.38 Mb, with a contig N50 of 1.37 Mb. Over 99% of the assembly was anchored to eight pseudochromosomes, with a scaffold N50 of 34.46 Mb. A total of 29,708 protein-coding genes were predicted from the genome and 46.85% (132.32 Mb) of the genome was annotated as repetitive sequence. Bud dormancy is influenced by chilling requirement in plum and partly controlled by DORMANCY ASSOCIATED MADS-box (DAM) genes. Six tandemly arrayed PsDAM genes were identified in the assembled genome. Sequence analysis of PsDAM6 in "Sanyueli" revealed the presence of large insertions in the intron and exon regions. Transcriptome analysis indicated that the expression of PsDAM6 in the dormant flower buds of "Sanyueli" was significantly lower than that in the dormant flower buds of the high chill requiring "Furongli" plum. In addition, PsDAM6 expression was repressed by chilling treatment. The genome sequence of "Sanyueli" plum provides a valuable resource for elucidating the molecular mechanisms responsible for the regulation of chilling requirements, and it is also useful for the identification of the genes involved in the control of other important agronomic traits and molecular breeding in plum.

Assembly of the Complete Mitochondrial Genome of Chinese Plum (Prunus salicina): Characterization of Genome Recombination and RNA Editing Sites.

Despite the significant progress that has been made in the genome sequencing of Prunus, this area of research has been lacking a systematic description of the mitochondrial genome of this genus for a long time. In this study, we assembled the mitochondrial genome of the Chinese plum (Prunus salicina) using Illumina and Oxford Nanopore sequencing data. The mitochondrial genome size of P. salicina was found to be 508,035 base pair (bp), which is the largest reported in the Rosaceae family to date, and P. salicina was shown to be 63,453 bp longer than sweet cherry (P. avium). The P. salicina mitochondrial genome contained 37 protein-coding genes (PCGs), 3 ribosomal RNA (rRNA) genes, and 16 transfer RNA (tRNA) genes. Two plastid-derived tRNA were identified. We also found two short repeats that captured the nad3 and nad6 genes and resulted in two copies. In addition, nine pairs of repeat sequences were identified as being involved in the mediation of genome recombination. This is crucial for the formation of subgenomic configurations. To characterize RNA editing sites, transcriptome data were used, and we identified 480 RNA editing sites in protein-coding sequences. Among them, the initiation codon of the nad1 gene confirmed that an RNA editing event occurred, and the genomic encoded ACG was edited as AUG in the transcript. Combined with previous reports on the chloroplast genome, our data complemented our understanding of the last part of the organelle genome of plum, which will facilitate our understanding of the evolution of organelle genomes.

Chromosome-scale genome assembly and population genomics provide insights into the adaptation, domestication, and flavonoid metabolism of Chinese plum.

Globally, commercialized plum cultivars are mostly diploid Chinese plums (Prunus salicina Lindl.), also known as Japanese plums, and are one of the most abundant and variable fruit tree species. To advance Prunus genomic research, we present a chromosome-scale P. salicina genome assembly, constructed using an integrated strategy that combines Illumina, Oxford Nanopore, and high-throughput chromosome conformation capture (Hi-C) sequencing. The high-quality genome assembly consists of a 318.6-Mb sequence (contig N50 length of 2.3 Mb) with eight pseudo-chromosomes. The expansion of the P. salicina genome is led by recent segmental duplications and a long terminal repeat burst of approximately 0.2 Mya. This resulted in a significant expansion of gene families associated with flavonoid metabolism and plant resistance, which impacted fruit flavor and increased species adaptability. Population structure and domestication history suggest that Chinese plum may have originated from South China and provides a domestication route with accompanying genomic variations. Selection sweep and genetic diversity analysis enabled the identification of several critical genes associated with flowering time, stress tolerance, and flavonoid metabolism, demonstrating the essential roles of related pathways during domestication. Furthermore, we reconstructed and exploited flavonoid-anthocyanin metabolism using multi-omics analysis in Chinese plum and proposed a complete metabolic pathway. Collectively, our results will facilitate further candidate gene discovery for important agronomic traits in Chinese plum and provide insights into future functional genomic studies and DNA-informed breeding.

Evaluation of the S-locus in Prunus domestica, characterization, phylogeny and 3D modelling.

Self-compatibility has become the primary objective of most prune (Prunus domestica) breeding programs in order to avoid the problems related to the gametophytic self-incompatibility (GSI) system present in this crop. GSI is typically under the control of a specific locus., known as the S-locus., which contains at least two genes. The first gene encodes glycoproteins with RNase activity in the pistils., and the second is an SFB gene expressed in the pollen. There is limited information on genetics of SI/SC in prune and in comparison., with other Prunus species, cloning., sequencing and discovery of different S-alleles is very scarce. Clear information about S-alleles can be used for molecular identification and characterization of the S-haplotypes. We determined the S-alleles of 36 cultivars and selections using primers that revealed 17 new alleles. In addition, our study describes for the first time the association and design of a molecular marker for self-compatibility in P. domestica. Our phylogenetic tree showed that the S-alleles are spread across the phylogeny, suggesting that like previous alleles detected in the Rosaceae., they were of trans-specific origin. We provide for the first time 3D models for the P. domestica SI RNase alleles as well as in other Prunus species, including P. salicina (Japanese plum), P. avium (cherry), P. armeniaca (apricot), P. cerasifera and P. spinosa.

Molecular and Evolutionary Characterization of Pollen S Determinant (SFB Alleles) in Four Diploid and Hexaploid Plum Species (Prunus spp.).

In more than 60 families of angiosperms, the self- and cross-fertilization is avoided through a complex widespread genetic system called self-incompatibility (SI). One of the major puzzling issues concerning the SI is the evolution of this system in species with complex polyploid genomes. Among plums, one of the first fruits species to attract human interest, polyploid species represent enormous genetic potential, which can be exploited in breeding programs. However, molecular studies in these species are very scarce due to the complexity of their genome. In order to study the SFB gene [the male component of gametophytic self-incompatibility system (GSI)] in plum species, 36 plum accessions belonging to diploid and hexaploid species were used. A total of 19 different alleles were identified; 1 of them was revealed after analyzing sequences. Peptide sequence analysis allowed identifying the five domains features of the SFB gene. Polymorphism analysis showed a subtle difference between domesticated and open pollinated Tunisian accessions and suggested a probable influence of the ploidy level. Divergence analysis between studied sequences showed that a new specificity may appear after 5.3% of divergence at synonymous sites between pairs of sequences in Prunus insititia, 6% in Prunus cerasifera, 8% and 9% in Prunus domestica and Prunus salicina respectively. Furthermore, sites under positive selection, the ones more likely to be responsible for specificity determination, were identified. A positive and significant Pearson correlation was found between the divergence between sequences, divergence time, fixed substitutions (MK test), and PSS number. These results supported the model assuming that functionally distinct proteins have arisen not as a result of chance fixation of neutral variants, but rather as a result of positive Darwinian selection. Further, the role that plays recombination can not be ruled out, since a rate of 0.08 recombination event per polymorphic sites was identified.

Effect of benzothiadiazole treatment on quality and anthocyanin biosynthesis in plum fruit during storage at ambient temperature.

BACKGROUND: Plums tend to experience a reduction in fruit quality due to ripening and they deteriorate quickly during storage at room temperature. Benzothiadiazole (BTH) is a plant elicitor capable of inducing disease resistance in many crops. In this study, the effect of BTH treatment on fruit ripening, fruit quality, and anthocyanin biosynthesis in 'Taoxingli' plum was investigated. RESULTS: The results showed that BTH treatment could accelerate fruit ripening without affecting the incidence of fruit decay or the shelf life. Benzothiadiazole treatment improved the quality and consumer acceptability of 'Taoxingli' plums during storage by increasing the sweetness, red color formation, and the concentration of healthy antioxidant compounds. The BTH treatment could also effectively promote the biosynthesis of anthocyanin by enhancing the enzyme activities of phenylalanine ammonia-lyase (PAL), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), and uridine diphosphate flavonoid 3-O-glucosyltransferase (UFGT) and up-regulating the gene expressions of PsPAL, PsCHI, PsDFR, PsANS, and PsUFGT during storage. CONCLUSION: Benzothiadiazole treatment could be a potential postharvest technology for improving fruit quality and consumer acceptability in harvested plum fruit. © 2020 Society of Chemical Industry.

Chromosome-level draft genome of a diploid plum (Prunus salicina).

BACKGROUND: Plums are one of the most economically important Rosaceae fruit crops and comprise dozens of species distributed across the world. Until now, only limited genomic information has been available for the genetic studies and breeding programs of plums. Prunus salicina, an important diploid plum species, plays a predominant role in modern commercial plum production. Here we selected P. salicina for whole-genome sequencing and present a chromosome-level genome assembly through the combination of Pacific Biosciences sequencing, Illumina sequencing, and Hi-C technology. FINDINGS: The assembly had a total size of 284.2 Mb, with contig N50 of 1.78 Mb and scaffold N50 of 32.32 Mb. A total of 96.56% of the assembled sequences were anchored onto 8 pseudochromosomes, and 24,448 protein-coding genes were identified. Phylogenetic analysis showed that P. salicina had a close relationship with Prunus mume and Prunus armeniaca, with P. salicina diverging from their common ancestor ∼9.05 million years ago. During P. salicina evolution 146 gene families were expanded, and some cell wall-related GO terms were significantly enriched. It was noteworthy that members of the DUF579 family, a new class involved in xylan biosynthesis, were significantly expanded in P. salicina, which provided new insight into the xylan metabolism in plums. CONCLUSIONS: We constructed the first high-quality chromosome-level plum genome using Pacific Biosciences, Illumina, and Hi-C technologies. This work provides a valuable resource for facilitating plum breeding programs and studying the genetic diversity mechanisms of plums and Prunus species.

Interspecific introgression and natural selection in the evolution of Japanese apricot (Prunus mume).

Domestication and population differentiation in crops involve considerable phenotypic changes. The logs of these evolutionary paths, including natural/artificial selection, can be found in the genomes of the current populations. However, these profiles have been little studied in tree crops, which have specific characters, such as long generation time and clonal propagation, maintaining high levels of heterozygosity. We conducted exon-targeted resequencing of 129 genomes in the genus Prunus, mainly Japanese apricot (Prunus mume), and apricot (Prunus armeniaca), plum (Prunus salicina), and peach (Prunus persica). Based on their genome-wide single-nucleotide polymorphisms merged with published resequencing data of 79 Chinese P. mume cultivars, we inferred complete and ongoing population differentiation in P. mume. Sliding window characterization of the indexes for genetic differentiation identified interspecific fragment introgressions between P. mume and related species (plum and apricot). These regions often exhibited strong selective sweeps formed in the paths of establishment or formation of substructures of P. mume, suggesting that P. mume has frequently imported advantageous genes from other species in the subgenus Prunus as adaptive evolution. These findings shed light on the complicated nature of adaptive evolution in a tree crop that has undergone interspecific exchange of genome fragments with natural/artificial selections.

Isolation of novel citrus and plum fruit promoters and their functional characterization for fruit biotechnology.

BACKGROUND: Promoters that confer expression in fruit tissues are important tools for genetic engineering of fruit quality traits, yet few fruit-specific promoters have been identified, particularly for citrus fruit development. RESULTS: In this study, we report five citrus fruit-specific/preferential promoters for genetic engineering. Additionally, we have characterized a novel fruit-preferential promoter from plum. Genes specifically expressed in fruit tissues were selected and their isolated promoter regions were fused with the GUSPlus reporter gene for evaluation in transgenic plants. Stable transformation in Micro-Tom tomato demonstrated that the candidate promoter regions exhibit differing levels of expression and with varying degrees of fruit specificity. CONCLUSIONS: Among the five candidate citrus promoters characterized in this study, the CitSEP promoter showed a fruit-specific expression pattern, while the CitWAX and CitJuSac promoters exhibited high fruit-preferential expression with strong activity in the fruit, weak activity in floral tissues and low or undetectable activity in other tissues. The CitVO1, CitUNK and PamMybA promoters, while exhibiting strong fruit-preferential expression, also showed consistent weak but detectable activity in leaves and other vegetative tissues. Use of these fruit specific/preferential promoters for genetic engineering can help with precise expression of beneficial genes and help with accurate prediction of the activity of new genes in host fruit plants.

Hydrogen peroxide generated by over-expression of cytosolic superoxide dismutase in transgenic plums enhances bacterial canker resistance and modulates plant defence responses.

H2O2 generated during the oxidative burst, plays important roles in plant defenses responses against pathogens. In this study we examined the role of H2O2 on bacterial canker resistance in transgenic plums over-expressing cytosolic superoxide dismutase. Three transgenic lines (C64, C66 and F12) with elevated levels of H2O2 accumulation showed enhanced resistance against bacterial canker disease caused by Pseudomonas syringae pv. syringae, when compared to the non-transformed control. Analysis of the expression of several genes involved in the plant-pathogen interaction showed that the expression of those involved in SA pathway (pr1 and npr1) and JA (lox3) were activated earlier and transiently in transgenic lines C66 and F12 when compared to the wild type. However, the expression of genes involved in anthocyanin synthesis (chi, chs, f3h, dfr, atcs, myb10) and ethylene (acs) was induced at very low levels whereas it was activated by the pathogen at exaggerated levels in the non-transformed line. These results suggest that resistance observed in transgenic lines over-producing H2O2 is correlated with an early and transient induction of defense genes associated with the SA and JA pathways and inhibition of gene expression associated with ethylene and anthocyanin biosynthesis.