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In the model species Arabidopsis thaliana, FRIGIDA (FRI) is a key regulator of flowering time and can inhibit flowering without vernalization. However, little information is available on the function in the Rosaceae family. Loquat (Eriobotrya japonica) belongs to the family Rosaceae and is a distinctive species, in which flowering can be induced without vernalization, followed by blooming in late-autumn or winter. To investigate the functional roles of FRI orthologs in this non-vernalization species, we isolated an FRI ortholog, dubbed as EjFRI, from loquat. Analyses of the phylogenetic tree and protein sequence alignment showed that EjFRI is assigned to eurosids I FRI lineage. Expression analysis revealed that the highest expression level of EjFRI was after flower initiation. Meanwhile, EjFRI was widely expressed in different tissues. Subcellular localization of EjFRI was only detected to be in the nucleus. Ectopic expression of EjFRI in wild-type Arabidopsis delayed flowering time. The expression levels of EjFRI in transgenic wild-type Arabidopsis were significantly higher than those of nontransgenic wild-type lines. However, the expression levels of AtFRI showed no significant difference between transgenic and nontransgenic wild-type lines. Furthermore, the upregulated AtFLC expression in the transgenic lines indicated that EjFRI functioned similarly to the AtFRI of the model plant Arabidopsis. Our study provides a foundation to further explore the characterization of EjFRI, and also contributes to illuminating the molecular mechanism about flowering in loquat.

Triploid loquat (Eriobotrya japonica (Thunb.) Lindl.) has greater vigor than their respective diploid and tetraploid parents, but the molecular basis of this triploid heterosis remains unclear. Recent studies have suggested that DNA methylation is involved in heterosis, which is a recognized method of suppressing gene expression. However, our previous studies revealed a trend of increased DNA methylation in triploid loquat hybrids compared to their parents. To elucidate the mechanism of triploid loquat heterosis, we investigated the levels and regulation of relative gene expression between hybrid and parental lines using RNA-Seq technology. We found that gene expression in the hybrid lines was down-regulated and gene expression analysis revealed that approximately 94.56 and 86.97% were expressed additively in triploid-A and triploid-B, respectively. Analyses of the allele-specific gene expression in the hybrids revealed significantly more Longquan-1 alleles were preferentially expressed in the two hybrid lines. Further analysis of cis- and trans-regulatory effects showed that gene expression variation between parental alleles is largely attributable to cis-regulatory variation in triploid loquat and analyses of genes belonging to cis-regulatory variation showed that 88-90% of cis genes contributed to an additive expression pattern. Taken together, our results suggest that gene expression variation in triploid loquat fundamentally cis-regulated may play a dominant role in triploid loquat heterosis.

Background: Aneuploidy, a condition caused by an imbalance between the relative dosages of chromosomes, generally produces a novel phenotype specific to the molecular karyotype. Few techniques are currently available for detecting the molecular karyotypes of aneuploids in plants. Results: Based on this imbalance in chromosome dosage, a new approach (referred to as 'SSR-qPCR') combining simple sequence repeat (SSR) markers and quantitative real-time PCR (qPCR) has been developed and utilized to detect some common aneuploids irrespective of heterozygosity. We screened 17 specific SSR markers covering all loquat linkage groups and redesigned 6 pairs of primers for SSR markers that can detect loquat chromosome aneuploidies. The SSR-qPCR detection results obtained for hybrid progeny and open-pollination progeny of triploid loquat showed diagnostic accuracies of 88.9% and 62.5%, respectively, compared with the chromosome preparation results. Conclusion: SSR-qPCR can detect loquat aneuploids and be used to construct the entire molecular karyotypes of aneuploid individuals. Therefore, this method offers a novel alternative for the detection of chromosome aneuploidies.

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Wild loquats (Eriobotrya japonica Lindl.) provide remarkable genetic resources for studying domestication and breeding improved varieties. Herein, we generate the first high-quality chromosome-level genome assembly of wild loquat, with the 45,791 predicted protein-coding genes. Analysis of comparative genomics indicated that loquat shared a common ancestor with apple and pear, and a recent whole-genome duplication event occurred in loquat prior to its divergence. Genome re-sequencing showed that the loquat germplasms were distinctly classified into wild and cultivated groups, and the commercial cultivars experienced allelic admixture. Compared with the cultivated loquats, the wild loquat genome showed very few selected genomic regions and had higher levels of genetic diversity. However, whole-genome scans of selective sweeps were mainly related to fruit quality, size, and flesh color during the domestication process. Large-scale transcriptome and metabolome analyses were further performed to identify the differentially expressed genes (DEGs) and differentially accumulated metabolites (DAMs) in the wild and cultivated loquats, at various fruit development stages. Unlike that in wild loquat, the key DEGs and DAMs involved in carbohydrate metabolism, plant hormone signal transduction, flavonoid biosynthesis, and carotenoid biosynthesis were significantly regulated in cultivated loquats during fruit development. These high-quality reference genome, re-sequencing, and large-scale transcriptome/metabolome data provide valuable resources for elucidating fruit domestication and molecular breeding in loquat.

Background Ploidy manipulation is effective in seedless loquat breeding, in which flesh color is a key agronomic and economic trait. Few techniques are currently available for detecting the genotypes of polyploids in plants, but this ability is essential for most genetic research and molecular breeding. Results We developed a system for genotyping by quantitative PCR (qPCR) that allowed flesh color genotyping in multiple tetraploid and triploid loquat varieties (lines). The analysis of 13 different ratios of DNA mixtures between two homozygous diploids (AA and aa) showed that the proportion of allele A has a high correlation (R2 = 0.9992) with parameter b [b = a1/(a1 + a2)], which is derived from the two normalized allele signals (a1 and a2) provided by qPCR. Cluster analysis and variance analysis from simulating triploid and tetraploid hybrids provided completely correct allelic configurations. Four genotypes (AAA, AAa, Aaa, aaa) were found in triploid loquats, and four (AAAA, AAAa, AAaa, Aaaa; absence of aaaa homozygotes) were found in tetraploid loquats. DNA markers analysis showed that the segregation of flesh color in all F1 hybrids conformed to Mendel's law. When tetraploid B431 was the female parent, more white-fleshed triploids occurred among the progeny. Conclusions qPCR can detect the flesh color genotypes of loquat polyploids and provides an alternative method for analyzing polyploid genotype and breeding, dose effects and allele-specific expression.

Black shank is a devastating disease in tobacco production worldwide, and resistance to this disease in Nicotiana plumbaginifolia Viv. tobacco is controlled by the Php gene. The position of the Php gene on chromosomes of N. plumbaginifolia remains unclear. The tobacco line TP‐1, which was derived from N. tabacum L. cv. Yunyan87 and N. plumbaginifolia, was used in the present study to help locate the Php gene on the N. plumbaginifolia chromosomes and understand the effects of the N. plumbaginifolia chromosome addition on the genomic expression of N. tabacum. An in vitro infection test verified that TP‐1 was as resistant to Phytophthora parasitica var. nicotianae race 0 as N. plumbaginifolia and more resistant than Yunyan87. According to cytogenetic analysis, TP‐1 was identified as a monosomic alien addition line and the alien chromosome was recognized as the ninth chromosome of N. plumbaginifolia through. Thus, the P. parasitica var. nicotianae race 0 resistance gene might be Php on chromosome 9 of N. plumbaginifolia. A superficial transcriptomic analysis showed that addition of the alien chromosome slightly affected the genomic expression of the host, with only 3.32% of differentially expressed genes (DEGs) identified between TP‐1 and Yunyan87. According to Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, most (9/13) genes classified as involved in plant–pathogen reactions, particularly five serine/threonine‐protein kinase genes, were upregulated in TP‐1. It is worth noting that flavonoid biosynthesis may be greatly downregulated in TP‐1: two genes were both downregulated in TP‐1 according to gene ontology analysis, and all 20 DEGs in the flavonoid biosynthesis pathway were downregulated in TP‐1 according to KEGG analysis.