Audrey Sebolt

Development and Evaluation of a Genome-Wide 6K SNP Array for Diploid Sweet Cherry and Tetraploid Sour Cherry

High-throughput genome scans are important tools for genetic studies and breeding applications. Here, a 6K SNP array for use with the Illumina Infinium® system was developed for diploid sweet cherry (Prunus avium) and allotetraploid sour cherry (P. cerasus). This effort was led by RosBREED, a community initiative to enable marker-assisted breeding for rosaceous crops. Next-generation sequencing in diverse breeding germplasm provided 25 billion basepairs (Gb) of cherry DNA sequence from which were identified genome-wide SNPs for sweet cherry and for the two sour cherry subgenomes derived from sweet cherry (avium subgenome) and P. fruticosa (fruticosa subgenome). Anchoring to the peach genome sequence, recently released by the International Peach Genome Initiative, predicted relative physical locations of the 1.9 million putative SNPs detected, preliminarily filtered to 368,943 SNPs. Further filtering was guided by results of a 144-SNP subset examined with the Illumina GoldenGate® assay on 160 accessions. A 6K Infinium® II array was designed with SNPs evenly spaced genetically across the sweet and sour cherry genomes. SNPs were developed for each sour cherry subgenome by using minor allele frequency in the sour cherry detection panel to enrich for subgenome-specific SNPs followed by targeting to either subgenome according to alleles observed in sweet cherry. The array was evaluated using panels of sweet (n = 269) and sour (n = 330) cherry breeding germplasm. Approximately one third of array SNPs were informative for each crop. A total of 1825 polymorphic SNPs were verified in sweet cherry, 13% of these originally developed for sour cherry. Allele dosage was resolved for 2058 polymorphic SNPs in sour cherry, one third of these being originally developed for sweet cherry. This publicly available genomics resource represents a significant advance in cherry genome-scanning capability that will accelerate marker-locus-trait association discovery, genome structure investigation, and genetic diversity assessment in this diploid-tetraploid crop group.

Cell number regulator genes in Prunus provide candidate genes for the control of fruit size in sweet and sour cherry

Striking increases in fruit size distinguish cultivated descendants from small-fruited wild progenitors for fleshy fruited species such as Solanum lycopersicum (tomato) and Prunus spp. (peach, cherry, plum, and apricot). The first fruit weight gene identified as a result of domestication and selection was the tomato FW2.2 gene. Members of the FW2.2 gene family in corn (Zea mays) have been named CNR (Cell Number Regulator) and two of them exert their effect on organ size by modulating cell number. Due to the critical roles of FW2.2/CNR genes in regulating cell number and organ size, this family provides an excellent source of candidates for fruit size genes in other domesticated species, such as those found in the Prunus genus. A total of 23 FW2.2/CNR family members were identified in the peach genome, spanning the eight Prunus chromosomes. Two of these CNRs were located within confidence intervals of major quantitative trait loci (QTL) previously discovered on linkage groups 2 and 6 in sweet cherry (Prunus avium), named PavCNR12 and PavCNR20, respectively. An analysis of haplotype, sequence, segregation and association with fruit size strongly supports a role of PavCNR12 in the sweet cherry linkage group 2 fruit size QTL, and this QTL is also likely present in sour cherry (P. cerasus). The finding that the increase in fleshy fruit size in both tomato and cherry associated with domestication may be due to changes in members of a common ancestral gene family supports the notion that similar phenotypic changes exhibited by independently domesticated taxa may have a common genetic basis.

Rosaceae conserved orthologous sequences marker polymorphism in sweet cherry germplasm and construction of a SNP-based map

The Rosaceae Conserved Orthologous Set (RosCOS) provides a gene-based genome-wide set of markers that have been used in comparative analyses of peach (Prunus persica), apple (Malus × domestica), and strawberry (Fragaria spp.). In order to extend the use of these RosCOS to sweet cherry (Prunus avium L.), we identified markers that are polymorphic in breeding germplasm. Ninety-five percent (595/627) of previously designed RosCOS primer pairs amplified a product in six sweet cherry cultivars predicted to represent the range of genetic diversity in breeding germplasm. A total of 45% (282/627) RosCOS were polymorphic among the six cultivars, and allele number ranged from 2 to 6, with a genome-wide mean of 2.35. A subset of 92 genome-wide single nucleotide polymorphisms (SNPs) corresponding to 76 RosCOS was analyzed in 36 founder accessions and progeny. The expected and observed heterozygosity suggested that 83% of the RosCOS were in Hardy–Weinberg equilibrium, implying that most RosCOS behave as neutral markers. Principal coordinate analysis (PCO) identified one wild accession and two Spanish landraces that clustered differently from the other accessions. The relatively high number of unique alleles found in the three differentially clustered selections suggested that their use as parents has potential to increase the genetic diversity in future US-bred cultivars. Of the 92 RosCOS SNPs, 81 SNPs that represented 68 genome-wide RosCOS segregated in four mapping populations. These RosCOS were mapped in four F1 populations, thereby greatly improving the genetic linkage map of sweet cherry.

Characterization of New Tart Cherry (Prunus cerasus L.): Selections Based on Fruit Quality, Total Anthocyanins, and Antioxidant Capacity

Tart cherries (Prunus cerasus L.) are rich in anthocyanins and possess high antioxidant activity. The objective of this study was to evaluate six Michigan tart cherry selections for different quality attributes; fruit weight, firmness, total soluble solids (TSS), titratable acidity, instrumental color parameters, total anthocyanins, and antioxidant capacity, determined as Oxygen Radical Absorbance Capacity (ORAC). Generally, significant (p < 0.01) differences were observed across tart cherry selections for fruit weight, firmness, total soluble solids, titratable acidity, and color values. As compared to 13.7°B for Montmorency (control), the TSS contents of all the tart cherry selections were significantly higher; ranging from 15.8 °B in selection 27–10(50) to 20.2°B in Erdi Jubileum. Fruit weight also showed significant differences, which were in the range of 3.95–8.17 g/fruit. In comparison to Montmorency, other tart cherry selections showed significantly higher titratable acidity (1.20–1.41% vs. 1.132%); higher anthocyanins (78.9–391.4 μg/g vs. 33.1 μg/g, as gallic acid equivalent); and higher ORAC values (up to 145.4% more). With respect to cost and better marketability, the results of this study could be useful for the cherry juice/concentrate industry.

Cherry leaf spot resistance in cherry (Prunus) is associated with a quantitative trait locus on linkage group 4 inherited from P. canescens

Cherry leaf spot (CLS), caused by the fungal pathogen Blumeriella jaapii (Rehm) Arx (telomorph Phloeosporella padi [Lib.] Arx), is a major disease in all humid cherry-growing regions worldwide causing leaf yellowing and defoliation. The diploid Prunus species, P. canescens, had previously been identified as a source of CLS resistance. Therefore, the objective of this study was to identify quantitative trait loci (QTL) for CLS resistance derived from P. canescens in both diploid sweet cherry (P. avium) and tetraploid sour cherry (P. cerasus). Because of the simpler genetics of diploid cherry, the initial investigation was done with P. canescens-derived materials from crosses with sweet cherry, followed by validation using P. canescens-derived plant materials from sour cherry. A major QTL controlling P. canescens-derived CLS resistance, named CLSR_G4, was identified on linkage group 4 in sweet cherry and validated in sour cherry. All CLS-resistant individuals had one P. canescens-derived allele for CLSR_G4. A second QTL may be necessary for CLS resistance as one-fifth–one-third of the progeny individuals with the P. canescens-derived allele for CLSR_G4 were susceptible.

A DNA test for fruit flesh color in tetraploid sour cherry (Prunus cerasus L.)

Fruit flesh color in tetraploid sour cherry (Prunus cerasus) is an important market-driven trait in the USA where the fruit from the dominant cultivar has brilliant red skin but clear/yellow flesh. This brilliant red color in the processed products differentiates products from sour cherries grown in the USA compared to those in Europe where the cultivars predominantly have dark purple-red flesh. In sweet cherry (P. avium), red skin and flesh colors were shown to be controlled by a major MYB10-associated locus. Sour cherry, which is derived from sweet cherry and ground cherry (P. fruticosa), also exhibits a range of flesh colors, but the genetic control of flesh color is not known. Our objectives were to test the hypothesis that the MYB10 locus controls flesh color in sour cherry and develop a predictive DNA test for dark purple-red flesh color. Pedigree-linked sour cherry plant materials were phenotyped for flesh color. Thirteen haplotypes for the sour cherry MYB10 region were distinguished based on markers scored from the use of the cherry 6K Infinium® II SNP array. Six haplotypes were significantly associated with variation in flesh color, supporting a role for MYB10 in controlling flesh color variation in sour cherry. A simple sequence repeat primer pair, designed from the peach genome sequence near MYB10, amplified a fragment that uniquely identified the haplotype that was associated with the darkest purple-red flesh color. This marker can be used for marker-assisted breeding to identify individuals that are predicted to have dark purple-red flesh.

Assessment of the Inheritance of Resistance and Tolerance in Cherry (Prunus sp.) to Blumeriella jaapii, the Causal Agent of Cherry Leaf Spot

Cherry leaf spot (CLS), caused by Blumeriella jaapii, is a serious fungal disease of sour cherry (Prunus cerasus) where ‘Montmorency’, the major cultivar grown in the United States, is highly susceptible. As many as 10 fungicide sprays can be required each growing season to combat this disease; therefore, developing CLS-resistant cultivars is a top breeding priority. Germplasm previously reported to be resistant or tolerant to CLS was acquired and incorporated into the sour cherry breeding program at Michigan State University (MSU) and includes three cherry species: sour cherry, sweet cherry (P. avium), and the wild species P. canescens. This study aimed to (i) compare the CLS disease progression profile of the susceptible cultivar ‘Montmorency’ with those of the resistant and tolerant germplasm, and (ii) gain an understanding of the inheritance of these resistance and tolerance traits by evaluating the host response of progeny individuals belonging to families derived from this germplasm. Significant differences were observed between the susceptible ‘Montmorency’ and the tolerant and resistant accessions in their response to CLS and its progression during the growing season. Evaluation of the CLS host responses of progeny individuals derived from this germplasm supported a dominant two gene model for P. canescens derived resistance and a recessive gene model for sweet cherry derived tolerance. These insights into disease progression and trait inheritance improve the efficiency and potential success of breeding sour cherry cultivars with durable resistance to CLS. This article is protected by copyright. All rights reserved.