Genetics

Apple (Malus pumila Mill.) is one of the most widespread and commercially important fruit crops worldwide. There is increasing interest in the genetics of tree architecture. Various dwarf genes have been described in the literature but few if any have been mapped and characterised. At the ARC-Bien Donné Experimental Farm are two young progenies, both derived from ‘McIntosh’, segregating for two novel dwarf traits, one monogenic and the other apparently digenic. These progenies are being scored with markers to map the traits and to identify potential candidate genes. And a transcriptomic approach will also be pursued to identify candidate genes. Combining the results will help clarify the molecular genetics and mechanisms behind these dwarf traits in apple. Tracing the genes in relevant pedigrees will help avoid raising progenies segregating for these traits in future.

High-throughput automated genotyping of plants requires a technology that is robust in the face of complex plant genomes, and cost-effective for processing large numbers of samples.

Plant genomes may be polyploid, comprised of high copy number sequences, and/or sub-population structural diversity. The Axiom® Genotyping Solution has three components that address these complexities. One, it uses the Axiom GT1 algorithm which dynamically adapts to cluster variation produced by plant genomes. Two, novel genotyping algorithms are used for markers in regions of high genomic complexity. Three, markers are sorted into relevant genomic classes, and the potentially most accurate and polymorphic markers are sorted into the PolyHighResolution class. The analysis workflow, algorithms and results are presented for allo-octoploid, cultivated strawberry samples that were genotyped with the IStraw90TM Axiom® Genotyping Array (in a 96-array layout) designed by the RosBREED project. Genotypes for SNPs in the PolyHighResolution class have greater than 99.8% reproducibility, and low Mendelian inheritance error rates.

Cost effective genotyping is enabled by the Axiom® 384-array layout, where 384 samples are simultaneously processed in a 384 microplate for analyzing up to 35,000 polyploid or 50,0000 diploid SNPs per sample. Markers that have been identified as highly performing, such as those in the PolyHighResolution class as identified by the Axiom® Genotyping Analysis on Axiom 96-array layout can be transferred to Axiom 384-array layout with 100% fidelity. The multi-species capability of the Axiom 384-array layout enables the design of a single array for markers from all rosaceous species that can be used in marker-assisted breeding.

The USDA-ARS National Plant Germplasm System has 33 species of wild Malus, many of which were acquired from plant explorations performed over the past 30 years. The phylogenetic relationships among these species were determined by chloroplast sequencing (1681 bp from four regions). Five primary clades of species were identified. Malus x domestica and the primary progenitor species, M. sieversii, M. orientalis, M. sylvestris, and M. prunifolia were localized within one of the two clades specific to species of Chinese origin. Malus fusca, native to the western US and Canada, also localized within this Chinese clade. The other three Malus clades originate from Taiwan (M. doumeri), southern Italy (M. florentina), and central/eastern North America (M. ioensis, M. coronaria, M. angustifolia). Genetic diversity and admixture of individuals within each clade and species was evaluated using nuclear microsatellite markers. Together these data provide the basis for integrating genetic structure information with species range distribution modeling approaches to prioritize novel collection sites.

Apple (Malus x domestica) production is #17 for agricultural products both in the U.S. and world with a value of more than $31 billion worldwide. This important perennial crop is expensive to produce, with high costs for land, labor and inputs. The industry is dominated by a relatively few number of scion cultivars and rootstocks, which increases its susceptibility to threats of new diseases, pests, and changing climate conditions. There are approximately 38 wild Malus species which are native to Asia, Europe and North America. Some of these wild species exhibit desirable resistance to biotic and abiotic stress, unique fruit quality, and useful rootstock traits. Apple is an Annex 1 crop and is covered under the multilateral system of The International Treaty (IT) on Plant Genetic Resources for Food and Agriculture. Countries that ratify the IT agree to make genetic resources and related information about these crops stored in their gene banks available to facilitate research and information exchange. Global Conservation Strategies have for many of the Annex 1 crops summarized the information available regarding gene bank collections, methods for conservation, and safe germplasm movement. This discussion will define the process of developing a global strategy for apple conservation. An expert committee will be chosen. A survey of global genebanks will be conducted. Global collections will be summarized and gaps will be discussed. The results will be summarized in recommendations for genebank standards for the management of apple genetic resources. Through the development of a Global Conservation Strategy for Apple, we aim to facilitate the flow of information and apple genetic resources internationally.

The genus Rubus (subfamily Rosoideae) contains an estimated 600-800 species distributed world-wide. Several of these, including black raspberry (Rubus occidentalis L.) are grown as crops. Since the early 1900s, the black raspberry industry in the United States has steadily declined due to lack of adapted, disease resistant cultivars. Renewed interest in production and breeding new cultivars has been fueled by news regarding potential health benefits of black raspberry bioactive compounds. We present a genetic linkage map comprised of simple sequence repeat (SSR) markers derived from both genomic and expressed sequence tag libraries, and single nucleotide polymorphic (SNP) markers derived from genotyping by sequencing (GBS). The map was constructed using 93 progeny of the full-sibling population ORUS 4305 (ORUS 3021-2 x ORUS 4153-1). The map consists of seven linkage groups representing the seven haploid chromosomes of this diploid species. The consensus map covers 613.1 cM with the longest linkage group spanning 101.7 cM with 103 markers (G1) and the shortest spanning 77.6 cM with 61 markers (G7). The construction of a densely populated genetic linkage map will be used for quantitative trait locus (QTL) mapping of economically important traits and for comparative genomic studies with other members of the Rosaceae.

Chile is the first peach exporter in the South Hemisphere and the fifth producer in the world. Chilean initiatives of peach breeding have been carried out, where the fruit quality and postharvest performance are the main goals for the new peach varieties. Breeding is a time consuming and costly process. For this reason, the development of genomic tools to support the early selection of genotypes is quite relevant to improve the efficiency of breeding programs. Thus, the aim of this work was to identify genome regions associated with fruit quality traits in peach using a QTL analysis. Harvesting date, soluble solids content (SSC), color, weight, firmness, titratable acidity and mealiness were evaluated. The ‘Venus’ x ‘Venus’ segregating population was genotyped with microsatellite markers and SNPs using 9K SNP array for peach. A linkage map was built with 1,820 markers, which were mapped in 8 linkage groups. The resulting map spanned a total distance of 382.9 cM with an average of 0.21 cM between adjacent markers. A QTL for titratable acidity was associated to chromosome 1, another QTL linked to cover skin color was associated to chromosome 3, and QTLs detected in chromosome 4 were related with weight, harvesting date, soluble solids contents and firmness. Finally, six QTLs were detected for mealiness, four of them located in chromosome 4 and two in chromosome 2. These results provide insights into the genetic determinants of quality traits in peach, but further work is required to identify and to validate candidate genes and polymorphisms. (This work was supported by Fondecyt 11121396, FONDAP CRG 15070009; Genoma G13I0005, Conicyt fellowship D-21120635 to ACE and Basal PFB-16).

Cultivated apple (Malus × domestica) is the most important fruit crop of the temperate regions worldwide. The accurate phenotypic and genetic characterization of apple genetic resources is essential for enlarging the genetic bases of breeding populations. Here we investigated the genetic diversity and structure of almost 2,700 European dessert apple accessions (old cultivars) originating from 12 germplasm collections located in 9 European countries, Russia, and Kyrgyzstan using 16 common SSR markers. Funding derived from both national and European (FruitBreedomics) projects. Priority was given to accessions chosen to be both diverse and representative of regional/national landraces. Genotyping made the identification and clarification of many redundancies, among accessions with different names but identical SSR profiles, possible both within and between collections. A total of 1,750 unique diploid genotypes (unique SSR profiles) were retained for further analyses. The average allelic diversity was very high (Na ~23), as were the observed and expected heterozygosities (0.81-0.83). The genetic structure of these accessions was studied by Factorial Component Analysis (FCA) and Structure software. A slight differentiation according to the geographic origin of the accessions (when known) was found (Fst = 0.021 +/-0.003), when subdividing the accessions into 3 large regions (West, South, North-East). Three groups (K=3) were also identified by Structure, but did not fully coincide with the pre-defined geographic regions. Parentage analyses made it possible to infer parents of several old cultivars. Overall, the genetic diversity was very high, but with a weak structure confirming large gene flow across Europe.

During the last decade, the high-density marker map based on the F2 progeny of the ‘Texas’ almond and ‘Earlygold’ peach (TxE) cross has been a resource for the Prunus scientific community. TxE has been used to identify markers for the construction of anchored linkage maps in crosses involving peach and other Prunus and mapping major genes and QTLs on them, for fast mapping of markers or candidate genes with the bin mapping approach, for the comparison of maps with other rosaceous crops, for physical map construction, and recently as one of the reference maps employed for the alignment of the peach genome assembly. Once the peach genome sequence has become available, a new and more powerful avenue for most of these applications is open. This communication will present some of the new ways in which this population, and others derived from the ‘Texas’ x ‘Earlygold’ hybrid, are currently being used for a better understanding of the Prunus genome and the genetics of characters of interest for plant breeding. This will include: a) the new maps of TxE and the first backcross progeny to peach (T1E population) based on the 9k IPSC peach SNP chip, b) the resequencing of individuals of T1E to find the precise positions of recombination breakpoints, and c) the use of these two populations to study the inheritance of a broad set of characters and to identify alleles of almond which may be valuable for peach improvement.

We used genotyping by sequencing (GBS), a low-cost, high-throughput sequencing technology to genotype 333 Prunus accessions, preserved at the Prunus collection of the National Clonal Germplasm Repository (NCGR) in Davis, California. The Prunus collection is the second largest in this genebank with more than 90 taxa and in excess of 1600 accessions of Prunus spp that includes almonds, apricots, cherries, peaches and plums. The accessions genotyped here consist of heirlooms (old cultivars never patented, or off patent), landraces, breeder’s lines, and wild relatives of the peach from all over the world. Majority of accessions belonged to Prunus persica (84%), with 10% of them being wild relatives (P. mira, P. davidiana, and P. ferganensis) and 6% categorized as hybrids between peach and other related species and Prunus spp. The method produced on average 1 million sequence reads per accession, with majority of the accessions having more than 500,000 reads. We identified 18,008 single-nucleotide polymorphism (SNP) markers, present in at least ≥ 80% of analyzed accessions distributed across the entire genome. These genomic data will serve as a resource for breeders seeking to develop peach cultivars that will meet the challenge of changing climates, markets, and horticultural practices. The use of these SNP markers for conservation, management and utilization of the NCGR collection as well as for genome-wide association studies (GWAS) in combination with phenotypic data available through Germplasm Resources Information Network (GRIN) will be discussed.

Abstract
Auxin has been regarded as the main signal molecule coordinating the growth and ripening of fruits in strawberry, the reference genomic system for Rosaceae. Functional information regarding the key regulators of auxin biosynthesis during both vegetative and reproductive development and environmental responses is scarce in strawberry. We have isolated and characterized whole YUCCA family genes encoding the key enzyme for auxin biosynthesis in both diploid strawberry (Fragaria vesca) and octoploid strawberries (F. × ananassa Duch.). Transcriptional profiling was performed in different tissues, fruits of different developmental stages, and leaves under different abiotic stresses. Functionality of YUCs was studied in transgenic F. vesca by over-expression or RNA interference, also in F. × ananassa by tobacco rattle virus (TRV)-induced gene silencing (VIGS) technique. The genetic study showed that FvYUC6 has important role in vegetative and reproductive development in woodland strawberry. VIGS experiment provided evidence that the gene FaYUC11, most homologous to AtYUC11, is crucial for receptacle enlargement in octoploid strawberry. Also, our work supports the interpretation that auxin biosynthesis system in strawberry holds flexible responses to developmental and environmental signals. These results will provide useful information for strawberry production to improve plant growth and fruit yield.