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Feminized hemp seed producers often use selfing to maintain a strain name; however, selfing may lead to inferior plants for cannabidiol (CBD) production. Using three different hemp strains as parents [Candida (CD-1), Dinamed CBD, and Abacus], two outcrosses [Candida (CD-1) × Abacus and Dinamed CBD × Candida (CD-1)] and one self-cross [Candida (CD-1) × Candida (CD-1)] were conducted to produce feminized seed. Progeny from the self-cross were significantly smaller and had less yield than outcrossed progeny. Selfed progeny were variegated and highly variable for total dry weight and floral dry weight. Discriminant analysis of principal components (DAPC) using amplified fragment length polymorphism (AFLP) separated the three progeny populations and showed that outcrossed populations clustered closer to the maternal parent, possibly the result of a maternal effect. Analysis of molecular variance (AMOVA) indicated that most variation (74.5%) was within populations, because the progeny from all three populations are half-siblings of each other. The selfed progeny population had lower expected heterozygosity (He = 0.085) than each of the outcrossed progeny populations (He ≈ 0.10). These results suggest that selfed progeny may demonstrate inbreeding depression resulting from enhanced expression of homozygous recessive traits. It may be beneficial for feminized seed producers to use outcrossing instead of selfing to generate feminized seed for CBD production.

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The genus Aronia Medik., also known as chokeberry, is a group of deciduous shrubs in the Rosaceae family, subtribe Pyrinae. The four commonly accepted species include A. arbutifolia (L.) Pers., red chokeberry; A. melanocarpa (Michx.) Elliott, black chokeberry; A. prunifolia (Marshall) Reheder, purple chokeberry; and A. mitschurinii (A.K. Skvortsov & Maitul). Wild and domesticated Aronia species are found as diploids, triploids, and tetraploids. Genetic improvement of polyploid Aronia genotypes has been limited by suspected apomixis, which may be widespread or distinct to tetraploids. The objectives of this study were to elucidate the reproductive mechanisms of Aronia species and reveal the occurrence of apomixis within the genus and along ploidy lines. Twentynine Aronia accessions [five A. melanocarpa (23), five A. melanocarpa (43), eight A. prunifolia (33), four A. prunifolia (43), six A. arbutifolia (43), and one A. mitschurinii (43)] were used in this study. Intra-accession variability was evaluated by growing out progeny from each open-pollinated maternal accession and comparing plant phenotypes, ploidy levels, and amplified fragment length polymorphism (AFLP) marker profiles between the progeny and maternal accession. Progeny of diploid and tetraploid maternal plants had ploidy levels identical to maternal plants, except for UC009 (A. melanocarpa, 23) which produced a mix of diploids and tetraploids. UC143 and UC149 (A. prunifolia, 33) produced all triploid offspring, whereas all other triploid accessions produced offspring with variable ploidy levels including 23, 33, 43, and 53. Pentaploid Aronia has not been previously reported. Diploid accessions produced significant AFLP genetic variation (0.68-0.78 Jaccard's similarity coefficient) in progeny, which is indicative of sexual reproduction. Seedlings from tetraploid accessions had very little AFLP genetic variation (0.93-0.98 Jaccard's similarity coefficient) in comparison with their maternal accession. The very limited genetic variation suggests the occurrence of limited diplosporous apomixis with one round of meiotic division in tetraploid progeny. Triploid accessions appear to reproduce sexually or apomictically, or both, depending on the individual. These results support our understanding of Aronia reproductive mechanisms and will help guide future breeding efforts of polyploid Aronia species.

Aronia is a group of deciduous fruiting shrubs, of the Rosaceae family, native to eastern North America. Interest in Aronia has increased because of the high levels of dietary antioxidants in Aronia fruits. Using Illumina RNA-seq transcriptome analysis, this study investigates the molecular mechanisms of polyphenol biosynthesis during Aronia fruit development. Six A. melanocarpa (diploid) accessions were collected at four fruit developmental stages. De novo assembly was performed with 341 million clean reads from 24 samples and assembled into 90,008 transcripts with an average length of 801 bp. The transcriptome had 96.1% complete according to Benchmarking Universal Single-Copy Orthologs (BUSCOs). The differentially expressed genes (DEGs) were identified in flavonoid biosynthetic and metabolic processes, pigment biosynthesis, carbohydrate metabolic processes, and polysaccharide metabolic processes based on significant Gene Ontology (GO) biological terms. The expression of ten anthocyanin biosynthetic genes showed significant up-regulation during fruit development according to the transcriptomic data, which was further confirmed using qRT-PCR expression analyses. Additionally, transcription factor genes were identified among the DEGs. Using a transient expression assay, we confirmed that AmMYB10 induces anthocyanin biosynthesis. The de novo transcriptome data provides a valuable resource for the understanding the molecular mechanisms of fruit anthocyanin biosynthesis in Aronia and species of the Rosaceae family.

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Intergeneric hybridization between Aronia and Pyrus may provide a pathway for developing novel fruit types with larger, sweeter fruits, while maintaining the high levels of biologically health-promoting compounds present in Aronia fruits. Here we describe a deleterious genetic incompatibility, known as hybrid necrosis or hybrid lethality, that occurs in intergeneric F1 hybrids of Aronia melanocarpa x Pyrus communis and ×Sorbaronia dippelii x Pyrus communis. Pollination experiments revealed that maternal A. melanocarpa and ×S. dippelii pistils are compatible with pollen from P. communis. Controlled pollinations using different mating combinations resulted in varying levels of fruit and seed set. Because every combination produced at least some viable seeds, prezygotic incompatibility does not appear to be present. We attempted to recover putative intergeneric progeny via either in vitro germination or in vitro shoot organogenesis from cotyledons. Progeny of putative hybrids from A. melanocarpa x P. communis only survived for a maximum of 14 days before succumbing to hybrid lethality. Regeneration of ×S. dippelii x P. communis was successful for two seedlings that have been maintained for an extended time in tissue culture. These two seedlings have leaf morphologies intermediate between the two parental genotypes. We also confirmed their hybrid status by using AFLPs and flow cytometry. Putative intergeneric hybrids were grown out ex vitro before showing symptoms of hybrid necrosis and dying after 3 months. Eventually micrografts failed, ultimately showing the same symptoms of hybrid necrosis. These results show that intergeneric hybridization is possible between Aronia and related genera in the Rosaceae, but there are postzygotic barriers to hybridity that can prevent the normal growth and development of the progeny.