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Among the Mediterranean horticultural landraces, garlic is one of the crops most threatened by genetic erosion. Due to its sexual sterility and to the incidence of seed-borne diseases, historical varieties have been widely replaced by commercial cultivars. In Catalonia, despite the historical relevance of the crop, solely the Belltall garlic landrace is cultivated for commercial purposes. To assess the genotypic and phenotypic diversity within the Belltall garlic, we evaluated sixteen local accessions and five recognized traditional and modern varieties as controls. Genetic analysis with SSR and InDel markers showed low genetic diversity within the Belltall population, grouping modern and traditional varieties separately. Farmers and consumers were involved in the definition of the landrace ideotype and classified the materials by means of projective mapping. Scant phenotypic diversity was found within the Belltall landrace, which is characterized by its color profile and the small size of bulb and cloves. The Belltall landrace grown outside its area of origin lost the distinctive quality signals that differentiate the landrace from the commercial cultivars (clove appearance), indicating that the high quality of the landrace is under genotype-by-environment effects (i.e. local adaptation). Moreover, the size of the Belltall sowing clove had a strong effect on the harvested bulb size. Our research represents a case study for the description of the variability within garlic landraces and an approach to quantify the phenomenon of local adaptation that currently drives their conservation.

Among the Mediterranean horticultural landraces, garlic is one of the crops most threatened by genetic erosion. Due to its sexual sterility and to the incidence of seed-borne diseases, historical varieties have been widely replaced by commercial cultivars. In Catalonia, despite the historical relevance of the crop, solely the Belltall garlic landrace is cultivated for commercial purposes. To assess the genotypic and phenotypic diversity within the Belltall garlic, we evaluated sixteen local accessions and five recognized traditional and modern varieties as controls. Genetic analysis with SSR and InDel markers showed low genetic diversity within the Belltall population, grouping modern and traditional varieties separately. Farmers and consumers were involved in the definition of the landrace ideotype and classified the materials by means of projective mapping. Scant phenotypic diversity was found within the Belltall landrace, which is characterized by its color profile and the small size of bulb and cloves. The Belltall landrace grown outside its area of origin lost the distinctive quality signals that differentiate the landrace from the commercial cultivars (clove appearance), indicating that the high quality of the landrace is under genotype-by-environment effects (i.e. local adaptation). Moreover, the size of the Belltall sowing clove had a strong effect on the harvested bulb size. Our research represents a case study for the description of the variability within garlic landraces and an approach to quantify the phenomenon of local adaptation that currently drives their conservation., This research was supported by the Ajuntament de Passanant i Belltall, the Diputació de Tarragona, the Catalan Government through the Pla de la Biodiversitat Cultivada (grant ARP152/21/000010), and the grant ACCIÓ -Eurecat (Project PRIV2020-PLANTOMICS), This research was supported by the Ajuntament de Passanant i Belltall, the Diputació de Tarragona, the Catalan Government through the Pla de la Biodiversitat Cultivada (grant ARP152/21/000010), and the grant ACCIÓ -Eurecat (Project PRIV2020-PLANTOMICS), Postprint (published version)

We sequenced the genome of the highly heterozygous almond Prunus dulcis cv. Texas combining short and long-read sequencing. We obtained a genome assembly totaling 227.6 Mb of the estimated 238 Mb almond genome size, of which 91% is anchored to eight pseudomolecules corresponding to its haploid chromosome complement, and annotated 27,969 protein-coding genes and 6,747 non-coding transcripts. By phylogenomic comparison with the genomes of 16 additional close and distant species we estimated that almond and peach (P. persica) diverged around 5.88 Mya. These two genomes are highly syntenic and show a high degree of sequence conservation (20 nucleotide substitutions/kb). However, they also exhibit a high number of presence/absence variants, many attributable to the movement of transposable elements (TEs). TEs have generated an important number of presence/absence variants between almond and peach, and we show that the recent history of TE movement seems markedly different between them. TEs may also be at the origin of important phenotypic differences between both species, and in particular, for the sweet kernel phenotype, a key agronomic and domestication character for almond. Here we show that in sweet almond cultivars, highly methylated TE insertions surround a gene involved in the biosynthesis of amygdalin, whose reduced expression has been correlated with the sweet almond phenotype. Altogether, our results suggest a key role of TEs in the recent history and diversification of almond and its close relative peach.

Peach [Prunus persica L. Batsch] is one of the major temperate fruit tree species, the commercial materials of which have a low level of genetic variability. Almond [P. dulcis (Mill) DA Webb], a close relative of peach cultivated for its kernels, has a much higher level of diversity. The species are inter-compatible and often produce fertile hybrids, almond being a possible source of new genes for peach that could provide biotic and abiotic stress tolerance traits. In this paper we describe the development of a collection of peach-almond introgression lines (ILs) having a single fragment of almond (cv. Texas) in the peach background (cv. Earlygold). Lines with few introgressions were selected with markers from successive generations from a "Texas"× "Earlygold"F1 hybrid, initially using a set of SSRs and later with the 18 k peach SNP chip, allowing for the final extraction of 67 lines, 39 with almond heterozygous introgressions covering 99% of the genome, and 28 with homozygous introgressions covering 83% of the genome. As a proof of concept, four major genes and four quantitative characters were examined in the selected ILs giving results generally consistent with previous information on the genetics of these characters. This collection is the first of its kind produced in a woody perennial species and promises to be a valuable tool for genetic analyses, including dissection of quantitative traits, positional cloning, epistasis and as prebreeding material to introgress almond genes of interest into the peach commercial gene pool.

Japanese plum is a diploid fruit tree species, member of the Rosaceae family, generated by hybridization of Prunus salicina with diverse Prunus species. Japanese plum cultivars show great variability for fruit skin and flesh color, which are both major objectives in plum breeding. Subsequently, molecular markers for early selection of these traits in breeding programs are highly desirable. Despite candidate genes for fruit color have been identified in several Rosaceae species, no validated markers have been described for Japanese plum yet. In Rosaceae family, MYB10 transcription factor has been described as the main gene determining anthocyanin pigment accumulation in fruits, which is responsible for red, purple and black coloration. In order to design a useful marker for marker-assisted selection (MAS), we have explored the variability of the MYB10 gene group in Japanese plum and its association with fruit color. A set of primers were designed targeting conserved MYB10 domains using sequences from peach genome. Primer combinations were tested using a reduced set of P. salicina accessions. The primer pair amplifying more polymorphic alleles was selected and used to genotype a collection of 78 Japanese plum cultivars. One allele, present in all 51 skin-colored accessions and absent in the rest, was found associated with anthocyanin accumulation. This dominant marker can effectively predict the presence or absence of fruit skin coloration caused by anthocyanin pigments.

The red to blue hue of plant organs is caused due to anthocyanins, which are water-soluble flavonoid pigments. The accumulation of these pigments is regulated by a complex of R2R3-MYB transcription factors (TFs), basic-helix-loop-helix (bHLH), and WD-repeat (WDR) proteins (MBW complex). In Rosaceae species, R2R3-MYBs, particularly MYB10 genes, are responsible for part of the natural variation in anthocyanin colors. Japanese plum cultivars, which are hybrids of Prunus salicina, have high variability in the color hue and pattern, going from yellow-green to red and purple-blue, probably as a result of the interspecific hybridization origin of the crop. Because of such variability, Japanese plum can be considered as an excellent model to study the color determination in Rosaceae fruit tree species. Here, we cloned and characterized the alleles of the PsMYB10 genes in the linkage group LG3 region where quantitative trait loci (QTLs) for the organ color have been mapped to other Prunus species. Allele segregation in biparental populations as well as in a panel of varieties, combined with the whole-genome sequence of two varieties with contrasting fruit color, allowed the organization of the MYB10 alleles into haplotypes. With the help of this strategy, alleles were assigned to genes and at least three copies of PsMYB10.1 were identified in some varieties. In total, we observed six haplotypes, which were able to characterize 91.36% of the cultivars. In addition, two alleles of PsMYB10.1 were found to be highly associated with anthocyanin and anthocyanin-less skin. Their expression during the fruit development confirms their role in the fruit skin coloration. Here, we provide a highly efficient molecular marker for the early selection of colored or non-colored fruits in Japanese plum breeding programs.

Loss of genetic variability is an increasing challenge in tree breeding programs due to the repeated use of a reduced number of founder genotypes. However, in almond, little is known about the genetic variability in current breeding stocks, although several cases of inbreeding depression have been reported. To gain insights into the genetic structure in modern breeding programs worldwide, marker-verified pedigree data of 220 almond cultivars and breeding selections were analyzed. Inbreeding coefficients, pairwise relatedness, and genetic contribution were calculated for these genotypes. The results reveal two mainstream breeding lines based on three cultivars: “Tuono”, “Cristomorto”, and “Nonpareil”. Descendants from “Tuono” or “Cristomorto” number 76 (sharing 34 descendants), while “Nonpareil” has 71 descendants. The mean inbreeding coefficient of the analyzed genotypes was 0.041, with 14 genotypes presenting a high inbreeding coefficient, over 0.250. Breeding programs from France, the USA, and Spain showed inbreeding coefficients of 0.075, 0.070, and 0.037, respectively. According to their genetic contribution, modern cultivars from Israel, France, the USA, Spain, and Australia trace back to a maximum of six main founding genotypes. Among the group of 65 genotypes carrying the Sf allele for self-compatibility, the mean relatedness coefficient was 0.125, with “Tuono” as the main founding genotype (24.7% of total genetic contribution). The results broaden our understanding about the tendencies followed in almond breeding over the last 50 years and will have a large impact into breeding decision-making process worldwide. Increasing current genetic variability is required in almond breeding programs to assure genetic gain and continuing breeding progress.

Peach is an important stone fruit in worldwide temperate regions. Peach breeding is a very active field, and many programs are starting to benefit from the genomic resources produced by international efforts that are allowing the implementation of marker-assisted breeding (MAB) in some of them. IRTA-ASF-FruitFutur peach breeding program started its activity in 2004 and the integration of marker-assisted selection (MAS) in 2010. Since then, several Mendelian traits have been included in the MAS pipeline and new breeding strategies based on molecular markers are being implemented. MAS is being used mainly for subacid fruit (D), flat fruit shape (S), slow ripening (Sr), highlighter fruit skin (H), Melting/Non melting flesh (M) and pollen sterility (Ps). Markers are also available for glabrous skin (G), flesh colour (Y), and brachytic dwarfism (Dw) and Stony hard (Sh). Marker-assisted introgresión (MAI), another MAB strategy, is being applied to efficiently introgress new genetic variability coming from other Prunus species. We stablished the proof of concept using a cross between peach and almond that we are now expanding using other Prunus species. In parallel, and thanks to the knowledge acquired about two genes, almond fruit type (Alf) and juicy (Jui), we are trying to produce peach fruit in almond trees by introgressing both genes from peach into the almond genetic background. We also are developing another breeding strategy, Resynthesis, to obtain individuals very similar to a given elite heterozygous genotype. Finally, to deal with traits related to the adaptation to climate change it is important to estimate GxE interactions. For that, within the EU PRIMA Freeclimb project, we are studying a collection of diverse peach cultivars that were established in different European countries following the same experimental design. These strategies open new roads for the implementation of MAB in peach breeding programs.

Inventorying, characterising and conserving on-farm ancient olive trees is a priority for safeguarding their genetic, natural and agricultural value and for protecting ancient genotypes threatened with extinction. In the “Taula del Sénia” (M-TdS) area (northeast Iberian Peninsula) a highly important cultural landscape has been preserved, in which the olive groves play an outstanding social and economic role: the ancient olive trees, sustained by many local farmers, constitute a living heritage and provide a clear example of High Nature Value (HNV). A total of 4526 ancient productive olive trees, with a trunk circumference (PBH) larger than 3.5 m, were inventoried and their spatial localization and biometric measurements were collected. 41 olive trees have shown the highest category in monumentality (PBH >8.1 m). The outstanding trees might be 634–1082 years old. The endocarp morphology of a representative sample of the most ancient trees from this settlement resulted in 14 different profiles. The ancient trees genotyped, through eight simple sequence repeat (SSR) markers, revealed 43 SSR profiles. The use of SSR enabled us to verify that most of the trees (98%) belong to the local cv. ‘Farga’, a male sterile variety with a rare chlorotype, only a few trees corresponded with other local varieties, ‘Morrut’, ‘Canetera’ and ‘Sevillenca’, and ten hitherto unidentified genotypes were distinguished, some with chloroplast lineages different from the ‘Farga’ type. The M-TdS area holds a unique living and exploitable heritage with the highest concentration of ancient olive trees worldwide. On-farm conservation of this germplasm by the community of local growers is enabling preservation of this important source of genetic variation, potentially holding traits of resilience and adaptation to adverse soil and climatic conditions, demonstrated by the survival of these trees over the centuries. Farmers have undertaken initiatives to valorize the olive oil deriving from these M-Td

This paper proposes a new breeding strategy, marker-assisted introgression (MAI), to obtain lines of perennial species with a single introgressed fragment from a compatible species two generations after the interspecific hybrid. MAI allows enrichment of the genome of a species with genes from a wild or exotic relative in a short timeframe and with an intermediate step that allows a first exploration of genes/QTLs that the donor species can provide to the target crop. This method has three phases: (1) creating a large backcross one (BC1) population to select, with markers, a reduced number of individuals (15–30, called the prIL set) with a low number of introgressions; (2) phenotyping the prIL set for the traits of interest and inferring the inheritance and map position of segregating major genes/QTLs based on the known genotypes of the prILs; and (3) advancing selected lines carrying the traits of interest to a next generation of backcross or selfing to obtain individuals with a single introgression in the background of the elite commercial germplasm. The proof of concept of this strategy was implemented by using peach as the recurrent species and almond as the donor. The whole process can be done in 9–10 years as the identification of the first line with one introgression was after 5 years (2006–2011), and 4–5 additional years are needed for phenotypic evaluation of selected lines. The expansion of this method to other perennial clonally propagated crops and to other species of Prunus compatible with peach is discussed.

Genetic analysis of a diverse set of 42 traits for flower (5), phenology (9), fruit quality (19), leaf (8) and disease resistance (1) was carried out in two interspecific almond × peach populations, an F2 (T × E) and a BC1 (T1E), from the cross between ‘Texas’ almond and ‘Earlygold’ peach. Traits related to flower, phenology, fruit quality, leaf morphology and resistance to powdery mildew were phenotyped over 3 years in two locations and studied for co-segregation with a large set of SNP and SSR markers. Three maps were used, one for the T × E and two for the T1E (T1E and E) population. Nine major genes were identified and mapped: anther color (Ag/ag and Ag2/ag2), flower color (Fc2/fc2), maturity date (MD/md), almond fruit type (almond vs. peach; Alf/alf), juiciness (Jui/jui), blood flesh (DBF2/dbf2), powdery mildew resistance (Vr3) and flower type (showy/non-showy; Sh/sh). These genes were often located in genome positions different from those for major genes for similar traits mapped before. Two of them explain fundamental aspects that define the fruit of peach with respect to that of almond: Alf and Jui, for its thick and juicy mesocarp, respectively. The genetics of quantitative traits was studied, and 32 QTLs were detected, with consistent behavior over the years. New alleles identified from almond for important traits such as red skin color, blood flesh, fruit weight and powdery mildew resistance may prove useful for the introduction of new variability into the peach gene pool used in commercial breeding programs.

Progeny of certain peach crosses include plants with slow ripening (SR) fruits that do not mature and remain on the tree after leaf fall. This character is determined by a single gene (Sr/sr) that segregated in the F1 population of the cross between ‘Belbinette’ and ‘Nectalady’ (Bb × Nl). Heterozygous (Srsr) in both parents, the recessive srsr homozygote resulted in SR individuals. This gene mapped to linkage group 4 in a map constructed with SSRs in BbxNl, at the same genomic region previously found to contain a gene for maturity date (MD). An allele of a SCAR marker (PSR2) developed from the candidate gene for MD (ppa008301m), and with high sequence similarity to NAC transcription factors, was diagnostic for the sr allele in the Bb × Nl progeny, a collection of 135 normally-ripening cultivars and breeding lines, and 467 offspring of different lines segregating for this character. We developed an SSR marker, CPP15636, at this location (39 kb from PSR2), which had one allele associated with the character although a few recombinant individuals were identified. Both PSR2 and CPP15636 can be used for early identification and selection of normally-ripening genotypes in breeding programs. Data of MD in the population studied indicates that sr could be an allele of the MD gene. Given that the MD locus is partly responsible for the variability of the maturity date, we suggest that the sr allele in heterozygosis could confer desirable properties of postharvest behavior or specific maturity dates and would be preferentially selected in peach breeding.

Peach (Prunus persica) and almond (Prunus dulcis) are two sexually compatible species that produce fertile offspring. Almond, a highly polymorphic species, is a potential source of new genes for peach that has a strongly eroded gene pool. Here we describe the genetics of a male sterile phenotype that segregated in two almond (‘Texas’) × peach (‘Earlygold’) progenies: an F2 (T×E) and a backcross one (T1E) to the ‘Earlygold’ parent. High-density maps were developed using a 9k peach SNP chip and 135 simple-sequence repeats. Three highly syntenic and collinear maps were obtained: one for the F2 (T×E) and two for the backcross, T1E (for the hybrid) and E (for ‘Earlygold’). A major reduction of recombination was observed in the interspecific maps (T×E and T1E) compared to the intraspecific parent (E). The E map also had extensive monomorphic genomic regions suggesting the presence of large DNA fragments identical by descent. Our data for the male sterility character were consistent with the existence of cytoplasmic male sterility, where individuals having the almond cytoplasm required the almond allele in at least one of two independent restorer genes, Rf1 and Rf2, to be fertile. The restorer genes were located in a 3.4 Mbp fragment of linkage group 2 (Rf1) and 1.4 Mbp of linkage group 6 (Rf2). Both fragments contained several genes coding for pentatricopeptide proteins, demonstrated to be responsible for restoring fertility in other species. The implications of these results for using almond as a source of novel variability in peach are discussed.

Peaches with low acidity are preferred in the market and this trait is usually selected in commercial breeding programs. A major gene (D/d) has been described for this character located on linkage group 5 of peach, where the low acid character is determined by the dominant D allele. In this paper, we analyze a collection of 231 varieties and 542 offspring to identify diagnostic markers for this character. The CPPCT040 single sequence repeat (SSR) is known to be tightly linked to D. We found that one of its alleles (193) is diagnostic for the subacid character and identified with high probability individuals with low acidity (titratable acidity <5.5 mg/l). The region around CPPCT040 was explored using 13 DNA fragments for a total of 5,297 bp, covering a length of 70.4 kbp of the peach genome. The sequenced fragments detected 19 single nucleotide polymorphisms (SNPs) and five indels. All subacid individuals shared a large haplotype (>24 kb) around CPPCT040, a region with higher than average SNPs between acid and subacid varieties. The CPPCT040 marker plus one of the SNPs identified (DH875) were used to genotype a collection of 542 seedlings, from different crosses expected to segregate for this character, which were phenotyped by tasting the fruit in the field. Data provided by both markers were always consistent and only 24 plants (4 %) did not fit the expectations. These markers and others that can be obtained from the haplotype identified can be readily used for marker-assisted selection in peach breeding.