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Background In water lily (Nymphaea) hybrid breeding, breeders often encounter non-viable seeds, which make it difficult to transfer desired or targeted genes of different Nymphaea germplasm. We found that pre-fertilization barriers were the main factor in the failure of the hybridization of Nymphaea. The mechanism of low compatibility between the pollen and stigma remains unclear; therefore, we studied the differences of stigma transcripts and proteomes at 0, 2, and 6 h after pollination (HAP). Moreover, some regulatory genes and functional proteins that may cause low pollen-pistil compatibility in Nymphaea were identified. Results RNA-seq was performed for three comparisons (2 vs 0 HAP, 6 vs 2 HAP, 6 vs 0 HAP), and the number of differentially expressed genes (DEGs) was 8789 (4680 were up-regulated), 6401 (3020 were up-regulated), and 11,284 (6148 were up-regulated), respectively. Using label-free analysis, 75 (2 vs 0 HAP) proteins (43 increased and 32 decreased), nine (6 vs 2 HAP) proteins (three increased and six decreased), and 90 (6 vs 0 HAP) proteins (52 increased and 38 decreased) were defined as differentially expressed proteins (DEPs). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that the DEGs and DEPs were mainly involved in cell wall organization or biogenesis, S-adenosylmethionine (SAM) metabolism, hydrogen peroxide decomposition and metabolism, reactive oxygen species (ROS) metabolism, secondary metabolism, secondary metabolite biosynthesis, and phenylpropanoid biosynthesis. Conclusions Our transcriptomic and proteomic analysis highlighted specific genes, incuding those in ROS metabolism, biosynthesis of flavonoids, SAM metabolism, cell wall organization or biogenesis and phenylpropanoid biosynthesis that warrant further study in investigations of the pollen-stigma interaction of water lily. This study strengthens our understanding of the mechanism of low pollen-pistil compatibility in Nymphaea at the molecular level, and provides a theoretical basis for overcoming the pre-fertilization barriers in Nymphaea in the future.

Although there has been a long history of cultivation and research on Nymphaea, the taxonomic relationships and evolutionary relationships among Nymphaea species remain controversial. The chloroplast (cp) genome can provide a new method to determine species origin, evolution, and phylogenetic relationships of Nymphaea. The complete cp genomes of seven Nymphaea species were sequenced by high-throughput sequencing technology, and the structural characteristics and phylogenetic relationships of cp genomes were analyzed. The cp genomes of seven species of Nymphaea had rich variation in large single-copy (LSC) and small single-copy (SSC) regions. A comparative cp genomic analysis of seven Nymphaea species showed that the cp gene sequence of Nymphaea was consistent, with no signs of reverse rearrangement. Using N. colorata as the reference sequence, a total of 8328 single nucleotide polymorphisms (SNPs) and 1579 insertions/deletions were obtained. The degree of variation of the cp genome of the seven Nymphaea species in rpoA–rpl20, rbcL–ndhC, ndhD–ndhF, and trnN-GUU–ndhA regions was relatively high. The ycf2 gene was positively selected. The phylogenetic analysis showed that the genus Nymphaea can be further divided into five subgenera.

In our earlier study, pre-fertilization barrier was one of the main factors causing the failure of the wide hybridization between Chrysanthemum grandiflorum ‘Yuhuaxingchen’ and C. nankingense, and seriously restricted the utilization of the excellent chrysanthemum germplasm, C. nankingense. In order to overcome the pre-fertilization barrier and obtain some hybrids, we carried out wide hybridization between them again in this study, and tentatively adopted three pollination techniques including mentor pollen, delayed pollination, and gibberellic acid treatment. The average number of pollen grains germinating on stigma at 24 h after pollination and the percentage of embryo were investigated with a temporary mount method and the technique of fluorescence microscopy. It was found that, compared with the control, delayed pollination, gibberellic acid treatment, and mentor pollen significantly increased the average number of pollen grains germinating on each stigma at 24 h after pollination by approximately 152, 235, and 182%, respectively. In addition, all the three pollination techniques significantly increased the percentage of normal embryos at different periods after artificial pollination in comparison to the control. Furthermore, seed sets of the crosses using the three special pollination techniques were 1.1, 1.4, and 2.0%, respectively, whereas we did not obtained any seeds in the control. Taken together, these results suggest that delayed pollination, gibberellic acid treatment, and mentor pollen are three useful pollination techniques for overcoming pre-fertilization barriers and increasing seed set in the wide hybridization between C. grandiflorum ‘Yuhuaxingchen’ and C. nankingense, and they may be widely applied for increasing breeding efficiency in other chrysanthemum wide hybridization facing similar pre-fertilization barriers.

Chrysanthemum grandiflorum ‘Yuhuaxingchen’ is an important commercial chrysanthemum cultivar with excellent ornamental quality but low drought tolerance, whereas C. indicum has exceptional drought tolerance. In our earlier study, many hybrid seeds between them were obtained through interspecific hybridization. In the present study, we selected six putative hybrid lines with most drought tolerance from all the hybrid lines by withholding water, indentified their facticity by chromosome counting, and then evaluated their drought tolerance through determining foliar electrolyte leakage (EL), contents of malondialdehyde (MDA) and proline, and plant survival rate after 20% polyethylene glycol 6000 treatment. It was found that 155 out of 282 seeds germinated and only 132 seedlings survived. In addition, chromosome and morphological analysis showed that the six putative hybrids were real hybrids and their morphological features were intermediate between their parents. Furthermore, the density of leaf epidermal hair, proline content, and plant survival rate were the highest in C. indicum and the lowest in C. grandiflorum among the six hybrids and their parents. In contrast, EL value and MDA content were the highest in C. grandiflorum and the lowest in C. indicum. These results suggest that some true hybrids with improved drought tolerance can be obtained through interspecific hybridization in chrysanthemum breeding. Therefore, interspecific hybridization between chrysanthemum cultivars and their wild species may become a promising way to improve their biotic and abiotic resistance in the future breeding.

Reproductive barriers often exist in the crosses between Dendranthema grandiflorum (Ramat.) Kitamura and its wild species and seriously result in low seed set, consequently reducing breeding efficiency. For the purpose of revealing the factors leading to low seed set, we investigated pollen viability, germination behavior of pollen grains on stigmas and embryo development in the crosses between D. grandiflorum and three wild species, D. nankingense (Nakai) Tzvel., D. indicum (L.) Des Moul. and D. zawadskii (Herb.) Tzvel. using technique of paraffin section, and light, fluorescence and scanning electron microscopy. The results indicated pollen viability of three wild species ranged from 20 to 25%. In the cross between D. grandiflorum and D. nankingense, very few pollen grains germinated on stigmas after pollination and most of them germinated abnormally. In addition, normal embryos were observed in 12% ovaries at 8 days after pollination and thereafter all the embryos aborted. In other two crosses, many pollen grains germinated on stigmas and pollen tubes penetrated stigmas normally after pollination. Moreover, normal embryos were observed in over 50% ovaries from 8 to 15 days after pollination in the cross between D. grandiflorum and D. indicum, and seed set was 59%. In the cross between D. grandiflorum and D. zawadskii, normal embryos were observed in 52% ovaries at 8 days after pollination. After that, however, most embryos degenerated and seed set was only 9%. These data suggest that pollen viability has no significant effects on seed set of the three crosses. Very few germinated pollen grains on stigmas and abnormal growth of most pollen tubes before fertilization, and embryo abortion are the main factors causing failure of the cross between D. grandiflorum and D. nankingense, whereas only embryo abortion is a main factor resulting in low seed set in the cross between D. grandiflorum and D. zawadskii and no barriers occur in the cross between D. grandiflorum and D. indicum.

Background Breeding programs for the water lotus (Nelumbo nucifera) are hampered by an inability to account for variation in seed set associated with crosses between different cultivars. We studied seed set in two reciprocal crosses between lotus cultivars (‘Guili’ × ‘Aijiangnan’ and ‘Molingqiuse’ × ‘Qinhuaiyanzhi') to obtain insights into factors that govern fecundity in these experimental hybrids. Pollen viability, stigma receptivity and embryo development were compared for each hybrid and reciprocal cross. Results Pollen viability of the individual cultivars ranged from 4.1% to 20.2%, with the highest level (>11.9%) for all cultivars observed from the earliest collected grains (05:00–06:00 a.m.). Stigmatic pollen germination peaked at 4 h after pollination and varied from 4.8 to 60.6 grains per stigma among the crosses. Production of normal embryos ranged from 7.6% to 58.8% at 1 d after pollination and from 0 to 25% by 11 d after pollination. Seed set in crosses (0.2–23.3%) was generally lower than in open-pollinated plants (8.4–26.5%). Similar to the germination results, seed set was substantially reduced in both reciprocal crosses. Conclusions These results suggested that poor pollen fertility, low stigma receptivity, and embryo abortion were responsible for the failure of the crosses ‘Molingqiuse’ × ‘Qinhuaiyanzhi’, ‘Qinhuaiyanzhi’ × ‘Molingqiuse’, and ‘Aijiangnan’ × ‘Guili’.