Pomological and Molecular Characterization of Apple Cultivars in the German Fruit Genebank.

Traditional varieties are a valuable tool in modern apple breeding. However, the use of synonyms and missing source documentation hinder an effective identification and conservation of relevant cultivars. During several projects, the authenticity and diversity of the apple cultivar collection of the German Fruit Genebank (GFG) was evaluated extensively. The trueness-to-type of 7890 apple trees was assessed on a pomological and molecular level. Pomological evaluations were performed by at least two experienced experts to identify the original cultivar names. On the molecular level, a set of 17 SSR markers was used to determine a unique genetic profile for each apple cultivar. The pomological and molecular characterization was expressed in terms of a comprehensive trueness-to-type criterion and the results were previously published as a well-curated dataset. In this study, the published dataset was analyzed to evaluate the quality and diversity of the apple collection of the GFG and highlight new findings based on phylogenetic and parentage analysis. The dataset contains 1404 unique genetic profiles corresponding to unambiguous cultivar names. Of these 1404 cultivars, 74% were assessed as true-to-type. The collection of diploid apple cultivars showed a high degree of expected heterozygosity (Hexp = 0.84). Genetic diversity in terms of year and location of origin was investigated with a STRUCTURE analysis. It was hypothesized that genetic diversity might decline overtime due to restrictive breeding programs. The results showed a shift dynamic between older and newer cultivars in one specific cluster, but no significant decrease in genetic diversity was observed in this study. Lastly, a parentage analysis was performed to check parental relationships based on historical research. Out of 128 parent-child trios, 110 trios resulted in significant relationships and reconfirmed the information from the literature. In some cases, the information from the literature was disproven. This analysis also allowed for readjusting the trueness-to-type criteria for previously undetermined cultivars. Overall, the importance of authenticity evaluations for gene bank cultivars was highlighted. Furthermore, the direct use of the dataset was shown by relevant investigations on the genetic diversity and structure of the apple cultivar collections of the GFG.

Identifying indicators of apple bud dormancy status by exposure to artificial forcing conditions.

Dormancy in temperate fruit trees is a mechanism of temporary growth suspension, which is vital for tree survival during winter. Studies on this phenomenon frequently employ scientific methods that aim to detect the timing of dormancy release. Dormancy release occurs when trees have been exposed to sufficient chill, allowing them to resume growth under conducive conditions. This study investigates dormancy dynamics in two apple (Malus × domestica Borkh.) cultivars, 'Nicoter' and 'Topaz', by sampling branches in an orchard over 14 weeks (2019 to 2020) and over 31 weeks (2021 to 2022) and subjecting them to a 42-day budbreak forcing period in a growth chamber. Temporal changes in budbreak percentages demonstrated dormancy progression in the studied apple cultivars and allowed the three main dormancy phases to be distinguished: paradormancy (summer dormancy), endodormancy (deep dormancy) and ecodormancy (spring dormancy), along with transition periods between them. Using these data, we explored the suitability of several alternative methods to determine endodormancy release. Tabuenca's test, which predicts dormancy release based on the differences in dry weights of buds with and without forcing, showed promise for this purpose. However, our data indicated a need for considerable adjustments and validation of this test. Bud weight and water content of buds in the orchard did not align with budbreak percentages under forcing conditions, rendering them unsuitable for determining endodormancy release in 'Nicoter' and 'Topaz'. Shoot growth cessation did not seem to be connected with either dormancy progression or dormancy depth of the studied cultivars, whereas leaf fall coincided with the beginning of the transition from endo- to ecodormancy. This work addresses methodological limitations in dormancy research and suggests considering the mean time to budbreak and budbreak synchrony as additional criteria to assess tree dormancy status.

Functional evidence on the involvement of the MADS-box gene MdDAM4 in bud dormancy regulation in apple.

Over the course of the year, temperate trees experience extremes in temperature and day length. In order to protect themselves from frost damage in winter, they enter a dormant state with no visible growth where all leaves are shed and buds are dormant. Also the young floral tissues need to withstand harsh winter conditions, as temperature fruit trees like apple develop their flower buds in the previous year of fruit development. So far, the genetic control of induction and release of dormancy is not fully understood. However, the transcription factor family of DORMANCY-Associated MADS-box (DAM) genes plays a major role in the control of winter dormancy. One of these genes is MdDAM4. This gene is expressed in the early phase of bud dormancy, but little is known about its function. Six transgenic apple lines were produced to study the function of MdDAM4 in apple. For plant transformation, the binary plasmid vector p9oN-35s-MdDAM4 was used that contains the coding sequence of MdDAM4 driven by the 35S promoter. Transgenicity of the lines was proven by PCR and southern hybridization. Based on siRNA sequencing and phenotypic observations, it was concluded that line M2024 overexpresses MdDAM4 whereas the gene is silenced in all other lines. Phenotyping of the transgenic lines provided evidence that the overexpression of MdDAM4 leads to an earlier induction and a later release of dormancy. Silencing this gene had exactly the opposite effects and thereby led to an increased duration of the vegetation period. Expression experiments revealed genes that were either potentially repressed or activated by MdDAM4. Among the potentially suppressed genes were several homologs of the cytokinin oxidase 5 (CKX5), five LOX homologs, and several expansins, which may indicate a link between MdDAM4 and the control of leaf senescence. Among the potentially activated genes is MdDAM1, which is in line with observed expression patterns during winter dormancy. MdDAM2, which shows little expression during endodormancy also appears to be activated by MdDAM4. Overall, this study provides experimental evidence with transgenic apple trees for MdDAM4 being an important regulator of the onset of bud dormancy in apple.

Synergistic approach of PCR-based fragment length analysis and amplicon deep sequencing reveals rich diversity of S-alleles in sweet cherries from the Caucasian region of origin.

Introduction: The self-incompatibility system in sweet cherry (Prunus avium L.) prevents fertilization with own or genetically related pollen, and is genetically determined by the multi-allelic S-locus. Therefore, determining S-alleles is crucial for plant breeding and fruit production, as it enables the selection of compatible combinations of S-genotypes for successful pollination. Methods: In this study, S-alleles were identified in a total of 260 genotypes from the Caucasian region, the species' center of origin. S-allele genotyping was conducted using PCR fragment length analysis with the standard marker PaConsI-F/R2 and reference genotypes, complemented by sequence analysis through amplicon deep sequencing. Results and discussion: The genotypes collected from Azerbaijan and Turkey exhibit a high allelic richness at the S-locus, particularly compared to modern sweet cherry cultivars worldwide. Nine previously undescribed S-alleles were identified and designated as S45, S46, S47, S48, S49, S50, S51, S52 and S53. Given the expected high diversity for other traits, this plant material represents a valuable resource for further breeding research and introgression of new traits in future breeding programs. Furthermore, our results underscore that fragment length alone may not be sufficient for unambiguous assignment of S-alleles due to minimal length differences between different alleles. To address this issue, an S-allele reference ladder was developed using the rich diversity for precise assignment of the S-alleles. This tool can be applied in future experiments as a robust and cost-effective method for accurate S-genotyping across different runs and laboratories. Additionally, several selected S-genotypes were planted in a trial field and will be maintained as an S-allele reference collection.

The structure of the tetraploid sour cherry 'Schattenmorelle' (Prunus cerasus L.) genome reveals insights into its segmental allopolyploid nature.

Sour cherry (Prunus cerasus L.) is an important allotetraploid cherry species that evolved in the Caspian Sea and Black Sea regions from a hybridization of the tetraploid ground cherry (Prunus fruticosa Pall.) and an unreduced pollen of the diploid sweet cherry (P. avium L.) ancestor. Details of when and where the evolution of this species occurred are unclear, as well as the effect of hybridization on the genome structure. To gain insight, the genome of the sour cherry cultivar 'Schattenmorelle' was sequenced using Illumina NovaSeqTM and Oxford Nanopore long-read technologies, resulting in a ~629-Mbp pseudomolecule reference genome. The genome could be separated into two subgenomes, with subgenome PceS_a originating from P. avium and subgenome PceS_f originating from P. fruticosa. The genome also showed size reduction compared to ancestral species and traces of homoeologous sequence exchanges throughout. Comparative analysis confirmed that the genome of sour cherry is segmental allotetraploid and evolved very recently in the past.

Rvi4 and Rvi15 are the same apple scab resistance genes.

The apple (Malus x domestica) scab (Venturia inaequalis) resistance genes Rvi4 and Rvi15 were mapped to a similar region on the top of linkage group 2 and both resistance genes elicit the same type of resistance reaction, i.e., a hypersensitive response; hence, it is suspected that the two genes may be the same. As the two resistance genes Rvi4 and Rvi15 are currently used in apple breeding, it is important to clarify whether the two resistance genes are the same or not. Several approaches were used to make this determination. First, the pedigree of the genotype GMAL 2473, the source of Rvi15, was reconstructed. GMAL 2473 was found to be an F1 of 'Russian seedling', the genotype, which is known to also be the source of Rvi4. Next, it was further demonstrated that 'Regia', a cultivar known to carry Rvi4 (and Rvi2), carries the same gene (Vr2-C), which was demonstrated to be the gene inducing Rvi15 resistance. Finally, it was shown that transgenic lines carrying Vr2-C are compatible with race 4 apple scab isolates. Taken all together, these results definitively demonstrate that Rvi4 and Rvi15 are the same resistance gene. For future studies, we suggest referring to this resistance with the first name that was assigned to this gene, namely Rvi4. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-023-01421-0.

Quantitative Trait Locus Mapping for Fire Blight Resistance in an F2 Population of Malus fusca MAL0045 Uncovers Novel Resistance Loci.

Several fire blight resistance loci in Malus genotypes map on different linkage groups (LGs) representing chromosomes of the domesticated apple. Prior genetics studies primarily focused on F1 populations. A strong resistance quantitative trait locus (QTL) explained up to 66% of phenotypic variance in an F1 progeny derived from crossing the highly resistant wild apple genotype Malus fusca MAL0045 and the highly susceptible apple cultivar 'Idared', which was previously mapped on LG10 (Mfu10) of MAL0045. Strains of the causative bacterial pathogen Erwinia amylovora, notably those that show a single nucleotide polymorphism in the avrRpt2EA effector protein sequence at position 156 (e.g., Ea3049), are more virulent and overcome some known fire blight resistance donors and their QTLs. However, MAL0045 is resistant to Ea3049 and Mfu10 is not overcome, but most of the F1 progeny were highly susceptible to this strain. This phenomenon led to the assumption that other putative resistance factors not segregating in the F1 progeny might be present in the genome of MAL0045. Here, we crossed F1 progeny together to obtain 135 F2 individuals. Facilitated by genotyping-by-sequencing and phenotypic assessments, we identified and mapped two novel resistance QTLs in these F2 individuals on LGs 4 and 15, which were not identified in the F1. To our knowledge, these are the first resistance QTLs mapped in F2 progeny in Malus. In addition, we report that neither MAL0045 nor Mfu10 is broken down by a highly aggressive U.S. strain, LA635, after analyses in the original F1 individuals. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Deciphering the Mechanism of Tolerance to Apple Replant Disease Using a Genetic Mapping Approach in a Malling 9 × M. × robusta 5 Population Identifies SNP Markers Linked to Candidate Genes.

Apple replant disease (ARD) is a worldwide economic risk in apple production. Although several studies have shown that the wild apple accession Malus × robusta 5 (Mr5) is ARD-tolerant, the genetics of this tolerance have not yet been elucidated. A genetic mapping approach with a biparental population derived from contrasting parents involving molecular markers provides a means for marker-assisted selection of genetically complex traits and for determining candidate genes. In this study, we crossed the ARD-tolerant wild apple accession Mr5 and the ARD-susceptible rootstock 'M9' and analyzed the resultant progeny for ARD tolerance. Hence, a high-density genetic map using a tunable genotyping-by-sequencing (tGBS) approach was established. A total of 4804 SNPs together with 77 SSR markers were included in the parental maps comprising 17 linkage groups. The phenotypic responses to ARD were evaluated for 106 offspring and classified by an ARD-susceptibility index (ASI). A Kruskal-Wallis test identified SNP markers and one SSR marker on linkage groups (LG) 6 and 2 that correlated with ARD tolerance. We found nine candidate genes linked with these markers, which may be associated with plant response to ARD. These candidate genes provide some insight into the defense mechanisms against ARD and should be studied in more detail.

Cryopreservation of Malus and Pyrus Wild Species in the 'Fruit Genebank' in Dresden-Pillnitz, Germany.

A unique and valuable diversity of the Malus and Pyrus wild species germplasm is maintained ex situ in field collections in the 'Fruit Genebank' in Dresden-Pillnitz, Germany. (1) Background: The establishment of a duplicate collection is necessary to preserve this material safely from abiotic and biotic stress factors. (2) Methods: Two different techniques, cryopreservation using dormant buds and PVS2 vitrification using in vitro shoot tips, were applied and compared. (3) Results: In Malus altogether 180 accessions belonging to 32 species were processed with an average recovery rate of 39% within ten years using the dormant bud method. Accessions, 116 in number, achieved the criterion of 40% recovery which was 64.44% of all accessions tested. In the case of Pyrus germplasm a total of 35 accessions of 21 species and both techniques for cryopreservation were tested. In the results of these experiments the PVS2 method led to better results compared to the dormant bud method. (4) Conclusions: In Malus the dormant bud technique will be the method of choice for the future to build up the duplicate collection. Further experiments using both techniques are planned in the future to make a final decision for Pyrus.

Genetic and Pomological Determination of the Trueness-to-Type of Sweet Cherry Cultivars in the German National Fruit Genebank.

Genebank collections preserve many old cultivars with ancient breeding history. However, often, cultivars with synonymous or incorrect names are maintained in multiple collections. Therefore, pomological and genetic characterization is an essential prerequisite for confirming trueness-to-type of cultivars in gene bank collections. In our study, 1442 single sweet cherry (Prunus avium L.) trees of the German Fruit Genebank were evaluated according to their trueness-to-type. For this purpose, pomological analysis was performed, in which the accessions were assigned totheir historical cultivar names. The pomological identifications were based on several historical reference sources, such as fruit references from historical cherry cultivar and fruit-stone collections, as well as historical pomological literature sources. In addition, the cherry trees were genetically analyzed for cultivar identity using 16 SSR markers. Based on pomological characterization and genetic analysis for the majority of the trees (86%), cultivar authenticity could be confirmed. Most markers were highly discriminating and powerful for cultivar identification. The cherry collection showed a high degree of genetic diversity, with an expected heterozygosity He = 0.67. Generally, high genetic admixture between cultivars of different geographic origin and year of origin was obtained after STRUCTURE analysis, demonstrating the extensive exchange of genetic information between cherry cultivars in the collection over time. However, the phylogenetic tree calculated by DARwin reflected the geographic origin of selected cherry cultivars. After parentage analysis with CERVUS, paternity could not be confirmed for three cultivars, indicating the necessity of further pedigree analysis for these cultivars. The results of our study underlined the general importance of evaluating the authenticity of cultivars in genebank collections based on genetic and pomological characterization.

Exploring epigenetic variation for breeding climate resilient apple crops.

Climate change with warmer winter and spring temperatures poses major challenges to apple fruit production. Long-term observations confirm the trend toward earlier flowering, which leads to an increased risk of frost damage. New breeding strategies are needed to generate cultivars that are able to stay largely unaffected by warmer temperatures. Recently, epigenetic variation has been proposed as a new resource for breeding purposes and seems suitable in principle for apple breeding. However, to serve as a new resource for apple breeding, it is necessary to clarify whether epigenetic variation can be induced by the environment, whether it can create phenotypic variation, and whether this variation is stable across generations. In this brief review, we summarize the impact of climate change on the timing of apple phenology, highlight how epigenetic variation can potentially support novel breeding strategies, and point out important features of epigenetic variation that are required for its application in breeding programs.

Recent Developments and Strategies for the Application of Agrobacterium-Mediated Transformation of Apple Malus × domestica Borkh.

Genetic transformation has become an important tool in plant genome research over the last three decades. This applies not only to model plants such as Arabidopsis thaliana but also increasingly to cultivated plants, where the establishment of transformation methods could still pose many problems. One of such plants is the apple (Malus spp.), the most important fruit of the temperate climate zone. Although the genetic transformation of apple using Agrobacterium tumefaciens has been possible since 1989, only a few research groups worldwide have successfully applied this technology, and efficiency remains poor. Nevertheless, there have been some developments, especially in recent years, which allowed for the expansion of the toolbox of breeders and breeding researchers. This review article attempts to summarize recent developments in the Agrobacterium-mediated transformation strategies of apple. In addition to the use of different tissues and media for transformation, agroinfiltration, as well as pre-transformation with a Baby boom transcription factor are notable successes that have improved transformation efficiency in apple. Further, we highlight targeted gene silencing applications. Besides the classical strategies of RNAi-based silencing by stable transformation with hairpin gene constructs, optimized protocols for virus-induced gene silencing (VIGS) and artificial micro RNAs (amiRNAs) have emerged as powerful technologies for silencing genes of interest. Success has also been achieved in establishing methods for targeted genome editing (GE). For example, it was recently possible for the first time to generate a homohistont GE line into which a biallelic mutation was specifically inserted in a target gene. In addition to these methods, which are primarily aimed at increasing transformation efficiency, improving the precision of genetic modification and reducing the time required, methods are also discussed in which genetically modified plants are used for breeding purposes. In particular, the current state of the rapid crop cycle breeding system and its applications will be presented.

Profiling of phytohormones in apple fruit and buds regarding their role as potential regulators of flower bud formation.

Apple (Malus × domestica Borkh.) cropping behavior, if not regulated, is often manifested by high yields of small-sized fruit in so called ON-years, which are usually followed by strongly reduced crop loads in OFF-years. Such cropping pattern is defined as biennial bearing and causes significant losses in apple production. The growth of apple fruit overlaps with the formation of flower buds, which remain dormant until the following spring. Earlier works proposed that some fruit-derived mobile compounds, as e.g., phytohormones, could suppress flower bud formation that thereby leads to biennial bearing. We addressed this hypothesis by analyzing 39 phytohormones in apple seeds, fruit flesh and by measuring phytohormone export from the fruits of the biennial bearing cultivar 'Fuji' and of the regular bearing cultivar 'Gala'. Moreover, we analyzed the same compounds in bourse buds from fruiting (ON-trees) and non-fruiting (OFF-trees) spurs of both apple cultivars over the period of flower bud formation. Our results showed that apple fruit exported at least 14 phytohormones including indole-3-acetic acid and gibberellin A3; however, their influence on flower bud formation was inconclusive. A gibberellin-like compound, which was detected exclusively in bourse buds, was significantly more abundant in bourse buds from ON-trees compared with OFF-trees. Cultivar differences were marked by the accumulation of trans-zeatin-O-glucoside in bourse buds of 'Gala' ON-trees, whereas the levels of this compound in 'Gala' OFF were significantly lower and comparable to those in 'Fuji' ON- and OFF-trees. Particular phytohormones including five cytokinin forms as well as abscisic acid and its degradation products had higher levels in bourse buds from OFF-trees compared with ON-trees and were therefore proposed as potential promotors of flower bud initiation. The work discusses regulatory roles of phytohormones in flower bud formation in apple based on the novel and to date most comprehensive phytohormone profiles of apple fruit and buds.

Identification of Candidate Genes Associated With Tolerance to Apple Replant Disease by Genome-Wide Transcriptome Analysis.

Apple replant disease (ARD) is a worldwide economic risk in apple cultivation for fruit tree nurseries and fruit growers. Several studies on the reaction of apple plants to ARD are documented but less is known about the genetic mechanisms behind this symptomatology. RNA-seq analysis is a powerful tool for revealing candidate genes that are involved in the molecular responses to biotic stresses in plants. The aim of our work was to find differentially expressed genes in response to ARD in Malus. For this, we compared transcriptome data of the rootstock 'M9' (susceptible) and the wild apple genotype M. ×robusta 5 (Mr5, tolerant) after cultivation in ARD soil and disinfected ARD soil, respectively. When comparing apple plantlets grown in ARD soil to those grown in disinfected ARD soil, 1,206 differentially expressed genes (DEGs) were identified based on a log2 fold change, (LFC) ≥ 1 for up- and ≤ -1 for downregulation (p < 0.05). Subsequent validation revealed a highly significant positive correlation (r = 0.91; p < 0.0001) between RNA-seq and RT-qPCR results indicating a high reliability of the RNA-seq data. PageMan analysis showed that transcripts of genes involved in gibberellic acid (GA) biosynthesis were significantly enriched in the DEG dataset. Most of these GA biosynthesis genes were associated with functions in cell wall stabilization. Further genes were related to detoxification processes. Genes of both groups were expressed significantly higher in Mr5, suggesting that the lower susceptibility to ARD in Mr5 is not due to a single mechanism. These findings contribute to a better insight into ARD response in susceptible and tolerant apple genotypes. However, future research is needed to identify the defense mechanisms, which are most effective for the plant to overcome ARD.

Proteomic differences in apple spur buds from high and non-cropping trees during floral initiation.

The cropping behavior of biennial apple (Malus ×domestica Borkh.) cultivars is irregular and often follows a biennial bearing pattern with 'On' years (high crop load and inhibited floral bud formation) followed by 'Off' years (little crop load and a promoted formation of floral buds). To study proteomic differences between floral and vegetative buds, trees of the strongly alternating cultivar 'Fuji' and the regular bearing cultivar 'Gala' were either completely thinned or not thinned at full bloom to establish two cropping treatments with no ('Off') or a high ('On') crop load, respectively. Student's t-Tests indicated significant differences of protein profiles in buds from 2-year old spurs from both treatments at each sampling date. Abundance patterns of protein clusters coincided with the onset of floral bud initiation and were most noticeable in buds from 'On' trees with a decreased abundance of key enzymes of the phenylpropanoid and flavonoid pathways and an increased abundance of histone deacetylase and ferritins. Furthermore, an increased abundance of proteins involved in histone and DNA methylation was found in the buds from 'Off' trees. This study presents the first large-scale, label-free proteomic profiling of floral and vegetative apple buds during the period of floral bud initiation. SIGNIFICANCE: Although several studies exist that address the complex developmental processes associated with the formation of floral buds in apple (Malus ×domestica Borkh.) at transcriptomic level, no data is available for explaining the difference between floral and vegetative buds or biennial and regular bearing cultivars on a proteomic level. This study presents the first large-scale, label-free proteomic profiling of floral and vegetative apple buds from the two cultivars 'Fuji' and 'Royal Gala' during the period of floral bud initiation and renders possible the development of suitable biomarkers for biennial bearing in apple.

Tracing CRISPR/Cas12a Mediated Genome Editing Events in Apple Using High-Throughput Genotyping by PCR Capillary Gel Electrophoresis.

The use of the novel CRISPR/Cas12a system is advantageous, as it expands the possibilities for genome editing (GE) applications due to its different features compared to the commonly used CRISPR/Cas9 system. In this work, the CRISPR/Cas12a system was applied for the first time to apple to investigate its general usability for GE applications. Efficient guide RNAs targeting different exons of the endogenous reporter gene MdPDS, whose disruption leads to the albino phenotype, were pre-selected by in vitro cleavage assays. A construct was transferred to apple encoding for a CRISPR/Cas12a system that simultaneously targets two loci in MdPDS. Using fluorescent PCR capillary electrophoresis and amplicon deep sequencing, all identified GE events of regenerated albino shoots were characterized as deletions. Large deletions between the two neighboring target sites were not observed. Furthermore, a chimeric composition of regenerates and shoots that exhibited multiple GE events was observed frequently. By comparing both analytical methods, it was shown that fluorescent PCR capillary gel electrophoresis is a sensitive high-throughput genotyping method that allows accurate predictions of the size and proportion of indel mutations for multiple loci simultaneously. Especially for species exhibiting high frequencies of chimerism, it can be recommended as a cost-effective method for efficient selection of homohistont GE lines.

The draft chromosome-level genome assembly of tetraploid ground cherry (Prunus fruticosa Pall.) from long reads.

Cherries are stone fruits and belong to the economically important plant family of Rosaceae with worldwide cultivation of different species. The ground cherry, Prunus fruticosa Pall., is an ancestor of cultivated sour cherry, an important tetraploid cherry species. Here, we present a long read chromosome-level draft genome assembly and related plastid sequences using the Oxford Nanopore Technology PromethION platform and R10.3 pore type. We generated a final consensus genome sequence of 366 Mb comprising eight chromosomes. The N50 scaffold was ~44 Mb with the longest chromosome being 66.5 Mb. The chloroplast and mitochondrial genomes were 158,217 bp and 383,281 bp long, which is in accordance with previously published plastid sequences. This is the first report of the genome of ground cherry (P. fruticosa) sequenced by long read technology only. The datasets obtained from this study provide a foundation for future breeding, molecular and evolutionary analysis in Prunus studies.

Corrigendum: Genes Involved in Stress Response and Especially in Phytoalexin Biosynthesis Are Upregulated in Four Malus Genotypes in Response to Apple Replant Disease.

[This corrects the article DOI: 10.3389/fpls.2019.01724.].

No Evidence of Unexpected Transgenic Insertions in T1190 - A Transgenic Apple Used in Rapid Cycle Breeding - Following Whole Genome Sequencing.

Rapid cycle breeding uses transgenic early flowering plants as crossbreed parents to facilitate the shortening of breeding programs for perennial crops with long-lasting juvenility. Rapid cycle breeding in apple was established using the transgenic genotype T1190 expressing the BpMADS4 gene of silver birch. In this study, the genomes of T1190 and its non-transgenic wild-type PinS (F1-offspring of 'Pinova' and 'Idared') were sequenced by Illumina short-read sequencing in two separate experiments resulting in a mean sequencing depth of 182× for T1190 and 167× for PinS. The sequencing revealed 8,450 reads, which contain sequences of ≥20 bp identical to the plant transformation vector. These reads were assembled into 125 contigs, which were examined to see whether they contained transgenic insertions or if they are not using a five-step procedure. The sequence of one contig represents the known T-DNA insertion on chromosome 4 of T1190. The sequences of the remaining contigs were either equally present in T1190 and PinS, their part with sequence identity to the vector was equally present in apple reference genomes, or they seem to result from endophytic contaminations rather than from additional transgenic insertions. Therefore, we conclude that the transgenic apple plant T1190 contains only one transgenic insertion, located on chromosome 4, and shows no further partial insertions of the transformation vector. Accession Numbers: JQ974028.1.

Evaluation of Scab and Mildew Resistance in the Gene Bank Collection of Apples in Dresden-Pillnitz.

A set of 680 apple cultivars from the Fruit Gene bank in Dresden Pillnitz was evaluated for the incidence of powdery mildew and scab in two consecutive years. The incidence of both scab and powdery mildew increased significantly in the second year. Sixty and 43 cultivars with very low incidence in both years of scab and powdery mildew, respectively, were analysed with molecular markers linked to known resistance genes. Thirty-five cultivars were identified to express alleles or combinations of alleles linked to Rvi2, Rvi4, Rvi6, Rvi13, Rvi14, or Rvi17. Twenty of them, modern as well as a few traditional cultivars known before the introduction or Rvi6 from Malus floribunda 821, amplified the 159 bp fragment of marker CH_Vf1 that is linked to Rvi6. Alleles linked to Pl1, Pld, or Plm were expressed from five cultivars resistant to powdery mildew. Eleven cultivars were identified to have very low susceptibility to both powdery mildew and scab. The information on resistance/susceptibility of fruit genetic resources towards economically important diseases is important for breeding and for replanting traditional cultivars. Furthermore, our work provides a well-defined basis for the discovery of undescribed, new scab, and powdery mildew resistance.