Genomewide selection for fruit quality traits in apple: breeding insights gained from prediction and postdiction.

Many fruit quality traits in apple (Malus domestica Borkh.) are controlled by multiple small-effect quantitative trait loci (QTLs). Genomewide selection (genomic selection) might be an effective breeding approach for highly quantitative traits in woody perennial crops with long generation times like apple. The goal of this study was to determine if genomewide prediction is an effective breeding approach for fruit quality traits in an apple scion breeding program. Representative apple scion breeding germplasm (nindividuals = 955), high-quality single nucleotide polymorphism (SNP) data (nSNPs = 977), and breeding program fruit quality trait data at harvest were analyzed. Breeding parents `Honeycrisp' and `Minneiska' were highly represented. Moderate to high predictive abilities were observed for most fruit quality traits at harvest. For example, when 25% random subsets of the germplasm set were used as training sets, mean predictive abilities ranged from 0.35 to 0.54 across traits. Trait, training and test sets, family size for within family prediction, and number of SNPs per chromosome affected model predictive ability. Inclusion of large-effect QTLs as fixed effects resulted in higher predictive abilities for some traits (e.g. percent red overcolor). Postdiction (i.e. retrospective) analyses demonstrated the impact of culling threshold on selection decisions. The results of this study demonstrate that genomewide selection is a useful breeding approach for certain fruit quality traits in apple.

A new method to reconstruct the direction of parent-offspring duo relationships using SNP array data and its demonstration on ancient and modern cultivars in the outcrossing species malus × domestica.

Unordered parent-offspring (PO) relationships are an outstanding issue in pedigree reconstruction studies. Resolution of the order of these relationships would expand the results, conclusions, and usefulness of such studies; however, no such PO order resolution (POR) tests currently exist. This study describes two such tests, demonstrated using SNP array data in the outcrossing species apple (Malus × domestica) on a PO relationship of known order ("Keepsake" as a parent of "Honeycrisp") and two PO relationships previously ordered only via provenance information. The first test, POR-1, tests whether some of the extended haplotypes deduced from homozygous SNP calls from one individual in an unordered PO duo are composed of recombinant haplotypes from accurately phased SNP genotypes from the second individual. If so, the first individual would be the offspring of the second individual, otherwise the opposite relationship would be present. The second test, POR-2, does not require phased SNP genotypes and uses similar logic as the POR-1 test, albeit in a different approach. The POR-1 and POR-2 tests determined the correct relationship between "Keepsake" and "Honeycrisp". The POR-2 test confirmed "Reinette Franche" as a parent of "Nonpareil" and "Brabant Bellefleur" as a parent of "Court Pendu Plat". The latter finding conflicted with the recorded provenance information, demonstrating the need for these tests. The successful demonstration of these tests suggests they can add insights to future pedigree reconstruction studies, though caveats, like extreme inbreeding or selfing, would need to be considered where relevant.

The use of shared haplotype length information for pedigree reconstruction in asexually propagated outbreeding crops, demonstrated for apple and sweet cherry.

Pedigree information is of fundamental importance in breeding programs and related genetics efforts. However, many individuals have unknown pedigrees. While methods to identify and confirm direct parent-offspring relationships are routine, those for other types of close relationships have yet to be effectively and widely implemented with plants, due to complications such as asexual propagation and extensive inbreeding. The objective of this study was to develop and demonstrate methods that support complex pedigree reconstruction via the total length of identical by state haplotypes (referred to in this study as "summed potential lengths of shared haplotypes", SPLoSH). A custom Python script, HapShared, was developed to generate SPLoSH data in apple and sweet cherry. HapShared was used to establish empirical distributions of SPLoSH data for known relationships in these crops. These distributions were then used to estimate previously unknown relationships. Case studies in each crop demonstrated various pedigree reconstruction scenarios using SPLoSH data. For cherry, a full-sib relationship was deduced for 'Emperor Francis, and 'Schmidt', a half-sib relationship for 'Van' and 'Windsor', and the paternal grandparents of 'Stella' were confirmed. For apple, 29 cultivars were found to share an unknown parent, the pedigree of the unknown parent of 'Cox's Pomona' was reconstructed, and 'Fameuse' was deduced to be a likely grandparent of 'McIntosh'. Key genetic resources that enabled this empirical study were large genome-wide SNP array datasets, integrated genetic maps, and previously identified pedigree relationships. Crops with similar resources are also expected to benefit from using HapShared for empowering pedigree reconstruction.

A DNA test for high incidence of soft scald and soggy breakdown postharvest disorders in Malus domestica Borkh.

The 'Honeycrisp' apple, an economically important cultivar and breeding parent, is prone to soft scald and soggy breakdown postharvest physiological disorders. Phenotypic evaluation of soft scald is time consuming and costly, making it an excellent target for DNA-informed breeding. The objective of this study was to develop a DNA test for a soft scald and soggy breakdown quantitative trait locus (QTL) on linkage group two (LG2) that was characterized in a previous study. 'Honeycrisp' is homozygous for the undesirable high disorder incidence haplotype (HDI) at this QTL. In this study, sixteen single nucleotide polymorphism markers were evaluated for their associations with the HDI haplotype in a set of 132 unique cultivars and important breeding parents. A DNA test was successfully developed utilizing KASP™ (Kompetitive Allele Specific PCR) chemistry to identify the number of HDI haplotypes in individuals. This test had a 100% accuracy for detecting homozygous unfavorable HDI individuals and has an expected 88% accuracy over all three haplotype copy groups across the evaluated germplasm. This DNA test is a promising tool for minimizing the chances of selecting individuals that exhibit high incidence of soft scald postharvest disorder in 'Honeycrisp'-related germplasm. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-021-01245-w.

Genetic analysis of stilbenoid profiles in grapevine stems reveals a major mQTL hotspot on chromosome 18 associated with disease-resistance motifs.

Grapevine (Vitis spp.) contains a wealth of phytochemicals that have received considerable attention due to health-promoting properties and biological activities as phytoalexins. To date, the genetic basis of the quantitative variations for these potentially beneficial compounds has been limited. Here, metabolic quantitative trait locus (mQTL) mapping was conducted using grapevine stems of a segregating F2 population. Metabolic profiling of grapevine stems was performed using liquid chromatography-high-resolution mass spectrometry (LC-HRMS), resulting in the detection of 1317 ions/features. In total, 19 of these features matched with literature-reported stilbenoid masses and were genetically mapped using a 1449-SNP linkage map and R/qtl software, resulting in the identification of four mQTLs. Two large-effect mQTLs that corresponded to a stilbenoid dimer and a trimer were mapped on chromosome 18, accounting for phenotypic variances of 29.0% and 38.4%. Functional annotations of these large-effect mQTLs on the VitisNet network database revealed a major hotspot of disease-resistance motifs on chromosome 18. This 2.8-Mbp region contains 48 genes with R-gene motifs, including variants of TIR, NBS, and LRR, that might potentially confer resistance to powdery mildew, downy mildew, or other pathogens. The locus also encompasses genes associated with flavonoid and biosynthetic pathways that are likely involved in the production of secondary metabolites, including phytoalexins. In addition, haplotype dosage effects of the five mQTLs further characterized the genomic regions for differential production of stilbenoids that can be applied in resistance breeding through manipulation of stilbenoid production in planta.

Correction: Genetics of zonal leaf chlorosis and genetic linkage to a major gene regulating skin anthocyanin production (MdMYB1) in the apple (Malus × domestica) cultivar Honeycrisp.

[This corrects the article DOI: 10.1371/journal.pone.0210611.].

Genetics of zonal leaf chlorosis and genetic linkage to a major gene regulating skin anthocyanin production (MdMYB1) in the apple (Malus × domestica) cultivar Honeycrisp.

'Honeycrisp' is a widely grown and acclaimed apple cultivar that is commonly used in breeding programs. It also has a well-documented tendency to develop the physiological disorder, zonal leaf chlorosis (ZLC). This disorder causes reduced photosynthetic capacity and is thought to be due to a problem with phloem loading, although the underlying genetics of the disorder have not previously been discerned. In order to understand the breeding implications of the disorder, six families with 'Honeycrisp' as a parent and one family with 'Honeycrisp' as both a maternal and paternal grandparent were evaluated for ZLC incidence over two years. One major quantitative trait locus (QTL) for ZLC incidence was identified on linkage group (LG) 9. A haplotype in 'Honeycrisp' that originated from grandparent 'Duchess of Oldenburg' was associated with increased ZLC incidence in offspring in both years and all families evaluated. The LG9 QTL was 5 to 10 cM from MdMYB1, which is a major gene regulating fruit skin anthocyanin production. 'Honeycrisp' is heterozygous for red fruit skin overcolor at MdMYB1. The 'Honeycrisp' haplotype at the LG9 QTL associated with increased ZLC is in linkage phase with the allele at MdMYB1 associated with red color. Selection for the red allele from 'Honeycrisp' at MdMYB1 will result in most offspring also inheriting the haplotype at the LG9 QTL associated with high ZLC. The occurrence of two copies of this haplotype was sub-lethal in seedlings of a family where both parents inherited both the red overcolor allele at MdMYB1 and the haplotype at the LG9 QTL associated with high ZLC. This is the first study to have identified a genetic component of ZLC with clear breeding implications.

Elucidation of the 'Honeycrisp' pedigree through haplotype analysis with a multi-family integrated SNP linkage map and a large apple (Malus×domestica) pedigree-connected SNP data set.

The apple (Malus×domestica) cultivar Honeycrisp has become important economically and as a breeding parent. An earlier study with SSR markers indicated the original recorded pedigree of 'Honeycrisp' was incorrect and 'Keepsake' was identified as one putative parent, the other being unknown. The objective of this study was to verify 'Keepsake' as a parent and identify and genetically describe the unknown parent and its grandparents. A multi-family based dense and high-quality integrated SNP map was created using the apple 8 K Illumina Infinium SNP array. This map was used alongside a large pedigree-connected data set from the RosBREED project to build extended SNP haplotypes and to identify pedigree relationships. 'Keepsake' was verified as one parent of 'Honeycrisp' and 'Duchess of Oldenburg' and 'Golden Delicious' were identified as grandparents through the unknown parent. Following this finding, siblings of 'Honeycrisp' were identified using the SNP data. Breeding records from several of these siblings suggested that the previously unreported parent is a University of Minnesota selection, MN1627. This selection is no longer available, but now is genetically described through imputed SNP haplotypes. We also present the mosaic grandparental composition of 'Honeycrisp' for each of its 17 chromosome pairs. This new pedigree and genetic information will be useful in future pedigree-based genetic studies to connect 'Honeycrisp' with other cultivars used widely in apple breeding programs. The created SNP linkage map will benefit future research using the data from the Illumina apple 8 and 20 K and Affymetrix 480 K SNP arrays.

Genetic dissection of powdery mildew resistance in interspecific half-sib grapevine families using SNP-based maps.

Quantitative trait locus (QTL) identification in perennial fruit crops is impeded largely by their lengthy generation time, resulting in costly and labor-intensive maintenance of breeding programs. In a grapevine (genus Vitis) breeding program, although experimental families are typically unreplicated, the genetic backgrounds may contain similar progenitors previously selected due to their contribution of favorable alleles. In this study, we investigated the utility of joint QTL identification provided by analyzing half-sib families. The genetic control of powdery mildew was studied using two half-sib F1 families, namely GE0711/1009 (MN1264 × MN1214; N = 147) and GE1025 (MN1264 × MN1246; N = 125) with multiple species in their ancestry. Maternal genetic maps consisting of 1077 and 1641 single nucleotide polymorphism (SNP) markers, respectively, were constructed using a pseudo-testcross strategy. Ratings of field resistance to powdery mildew were obtained based on whole-plant evaluation of disease severity. This 2-year analysis uncovered two QTLs that were validated on a consensus map in these half-sib families with improved precision relative to the parental maps. Examination of haplotype combinations based on the two QTL regions identified strong association of haplotypes inherited from 'Seyval blanc', through MN1264, with powdery mildew resistance. This investigation also encompassed the use of microsatellite markers to establish a correlation between 206-bp (UDV-015b) and 357-bp (VViv67) fragment sizes with resistance-carrying haplotypes. Our work is one of the first reports in grapevine demonstrating the use of SNP-based maps and haplotypes for QTL identification and tagging of powdery mildew resistance in half-sib families.

Genomic ancestry estimation quantifies use of wild species in grape breeding.

BACKGROUND: Grapes are one of the world's most valuable crops and most are made into wine. Grapes belong to the genus Vitis, which includes over 60 inter-fertile species. The most common grape cultivars derive their entire ancestry from the species Vitis vinifera, but wild relatives have also been exploited to create hybrid cultivars, often with increased disease resistance. RESULTS: We evaluate the genetic ancestry of some of the most widely grown commercial hybrids from North America and Europe. Using genotyping-by-sequencing (GBS), we generated 2482 SNPs and 56 indels from 7 wild Vitis, 7 V. vinifera, and 64 hybrid cultivars. We used a principal component analysis (PCA) based ancestry estimation procedure and verified its accuracy with both empirical and simulated data. V. vinifera ancestry ranged from 11 % to 76 % across hybrids studied. Approximately one third (22/64) of the hybrids have ancestry estimates consistent with F1 hybridization: they derive half of their ancestry from wild Vitis and half from V. vinifera. CONCLUSIONS: Our results suggest that hybrid grape breeding is in its infancy. The distribution of V. vinifera ancestry across hybrids also suggests that backcrosses to wild Vitis species have been more frequent than backcrosses to V. vinifera during hybrid grape breeding. This pattern is unusual in crop breeding, as it is most common to repeatedly backcross to elite, or domesticated, germplasm. We anticipate our method can be extended to facilitate marker-assisted selection in order to introgress beneficial wild Vitis traits, while allowing for offspring with the highest V. vinifera content to be selected at the seedling stage.

Next Generation Mapping of Enological Traits in an F2 Interspecific Grapevine Hybrid Family.

In winegrapes (Vitis spp.), fruit quality traits such as berry color, total soluble solids content (SS), malic acid content (MA), and yeast assimilable nitrogen (YAN) affect fermentation or wine quality, and are important traits in selecting new hybrid winegrape cultivars. Given the high genetic diversity and heterozygosity of Vitis species and their tendency to exhibit inbreeding depression, linkage map construction and quantitative trait locus (QTL) mapping has relied on F1 families with the use of simple sequence repeat (SSR) and other markers. This study presents the construction of a genetic map by single nucleotide polymorphisms identified through genotyping-by-sequencing (GBS) technology in an F2 mapping family of 424 progeny derived from a cross between the wild species V. riparia Michx. and the interspecific hybrid winegrape cultivar, 'Seyval'. The resulting map has 1449 markers spanning 2424 cM in genetic length across 19 linkage groups, covering 95% of the genome with an average distance between markers of 1.67 cM. Compared to an SSR map previously developed for this F2 family, these results represent an improved map covering a greater portion of the genome with higher marker density. The accuracy of the map was validated using the well-studied trait berry color. QTL affecting YAN, MA and SS related traits were detected. A joint MA and SS QTL spans a region with candidate genes involved in the malate metabolism pathway. We present an analytical pipeline for calling intercross GBS markers and a high-density linkage map for a large F2 family of the highly heterozygous Vitis genus. This study serves as a model for further genetic investigations of the molecular basis of additional unique characters of North American hybrid wine cultivars and to enhance the breeding process by marker-assisted selection. The GBS protocols for identifying intercross markers developed in this study can be adapted for other heterozygous species.

Changes in fruit antioxidant activity among blueberry cultivars during cold-temperature storage.

Antioxidant activity, total phenolic content, anthocyanin content, and six other fruit characters including titratable acid concentration, soluble solids, firmness, and percentage of bruised berries were determined for nine blueberry (Vaccinium L. sp.) cultivars at harvest and at various postharvest intervals after storage at 5 degrees C. Berries from MSU-58, Brigitta, and Legacy stored successfully for 7 weeks, Bluegold stored for 3-5 weeks, Bluecrop, Elliott, and Nelson stored for 3 weeks, and Jersey and Little Giant stored for fewer than 3 weeks. During the time they retained marketable quality, one cultivar (MSU-58) demonstrated a 29% increase in antioxidant activity. None of the cultivars showed a significant decrease from the harvest antioxidant activity value during storage. Antioxidant activity, total phenolic content, and anthocyanin content were strongly correlated with each other (r = 0.87-0.99, P < 0.01). All three parameters were moderately correlated with soluble solids (r = 0.47, P < or =0.05; r = 0.44, P < or = 0.05; and r = 0.64, P < or = 0.01, respectively), and antioxidant activity and total phenolic content were both moderately correlated with pH (r = 0.53 and 0.49, respectively; P < or = 0.05). However, antioxidant activity, total phenolic content, and anthocyanin content showed no correlation with firmness, percent severely bruised berries, or weight loss. Antioxidant activity and total phenolic content at harvest both correlated with titratable acidity at harvest (r = 0.68, P < or = 0.05 and r = 0.70, P < or = 0.05, respectively) on a cultivar mean basis. Berries from Elliott were also harvested from plants at two levels of bush ripeness (30-50% and 60-80% ripe berries on plants) and separated into three fruit maturity classes on the basis of percent blue color. The level of bush ripeness had no significant effect on antioxidant activity, total phenolic content, or anthocyanin content; however, fruit maturity had a significant effect on antioxidant activity, total phenolic content, and anthocyanin content, and bush ripeness x fruit maturity interactions were significant for these three traits. Berries with 50-75% blue coloration harvested from bushes with 60-80% mature fruit showed a significant increase in antioxidant activity, total phenolic content, and anthocyanin content during the first 3 weeks in storage. Our results demonstrate that increases in antioxidant activity, total phenolic content, and anthocyanin content may occur in the blueberry during cold storage and are cultivar-dependent. The increases that occur in immature fruit, such as in Elliott, may be advantageous for producers who wish to delay marketing of the fruit.