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Alternative splicing (AS) selects different alternative splice sites and produces a variety of transcripts with different exon/intron combinations, which may result in multiple protein isoforms. The splicing signals include cis-elements and RNA structures; however, the mechanisms of AS regulation in plants have yet to be elucidated. Previous studies have shown that in Platanus acerifolia, the FLOWERING LOCUS T (FT) homolog PaFT has a unique and complex AS pattern, in which most of the splice forms of PaFT involve the first and/or second intron, and the FD homolog PaFDL1 produces two transcripts via AS, whereas the other FT homolog PaFTL is not regulated by AS. In this study, the regulatory mechanism of the AS of PaFT was demonstrated to be conserved in different plant species. To define the distribution of the AS regulatory signals, the intron-swap, site-directed mutagenesis of alternative splice sites, and deletion experiment were performed. For the PaFT gene, all the signals that regulate the AS of the first intron were located within this intron, while the usage of the first alternative splice site in the second intron was determined by the first intron. Meanwhile, the AS of PaFDL1 might be co-regulated by exons and the first intron. Additionally, the first alternative splice site and adjacent region in PaFT intron 1 might contain cis-elements and/or RNA structures that affect the use of the other sites. This study had provided a deeper insight into the distribution of AS signals in plants, namely the AS signals of different splice sites might exist in the intron where the sites were present, and might also be distributed in exons or other introns.

Members of AP1/FUL subfamily genes play an essential role in the regulation of floral meristem transition, floral organ identity, and fruit ripping. At present, there have been insufficient studies to explain the function of the AP1/FUL-like subfamily genes in Asteraceae. Here, we cloned two euAP1 clade genes TeAP1-1 and TeAP1-2, and three euFUL clade genes TeFUL1, TeFUL2, and TeFUL3 from marigold (Tagetes erecta L.). Expression profile analysis demonstrated that TeAP1-1 and TeAP1-2 were mainly expressed in receptacles, sepals, petals, and ovules. TeFUL1 and TeFUL3 were expressed in flower buds, stems, and leaves, as well as reproductive tissues, while TeFUL2 was mainly expressed in flower buds and vegetative tissues. Overexpression of TeAP1-2 or TeFUL2 in Arabidopsis resulted in early flowering, implying that these two genes might regulate the floral transition. Yeast two-hybrid analysis indicated that TeAP1/FUL proteins only interacted with TeSEP proteins to form heterodimers and that TeFUL2 could also form a homodimer. In general, TeAP1-1 and TeAP1-2 might play a conserved role in regulating sepal and petal identity, similar to the functions of MADS-box class A genes, while TeFUL genes might display divergent functions. This study provides a theoretical basis for the study of AP1/FUL-like genes in Asteraceae species.

London plane tree is widely grown as a landscaping and street tree, but the release of its trichomes creates a serious air-borne pollution problem. Identifying the key genes that regulate the development of trichomes is, therefore, an important tool for the molecular breeding of

International audience; Dianthus chinensis is widely used as a landscaping plant in rock gardens and has high commercial value. However, few studies of this plant's regeneration and genetic transformation have been reported. In this study, cotyledons of D. chinensis were used as explants for regeneration and infection with Agrobacterium. The results indicated that the efficiency of regeneration was markedly improved by using 1-naphthylcetic acid (NAA). The cotyledons were cocultured with Agrobacterium tumefaciens EHA105 containing pK7GWIWG2-GFP plasmids for 15-20 min. Transgenic regenerated plants were obtained by kanamycin (80 mg.L-1) screening. qRT-PCR results confirmed the expression of GFP and the DcIPT1 (which is related to cell division) gene in the transgenic DcIPT1 plants of D. chinensis. The results show that the proliferation ability of transgenic DcIPT1 plants is enhanced. Overall, we report the successful establishment of a reliable and efficient method for cotyledon transformation and plant regeneration in D. chinensis based on the combination of infection with Agrobacterium and plant regeneration. This work will contribute to functional gene research and genetic improvement of Dianthus species plants.

The phosphatidylethanolamine-binding protein (PEBP) gene family exists in all eukaryote kingdoms, with three subfamilies: FT (FLOWERING LOCUS T)-like, TFL1 (TERMINAL FLOWER 1)-like, and MFT (MOTHER OF FT AND TFL1)-like. FT genes promote flowering, TFL1 genes act as a repressor of the floral transition, and MFT genes have functions in flowering promotion and regulating seed germination. We identified and characterized orthologs of the Arabidopsis FT/TFL1 gene family in petunia to elucidate their expression patterns and evolution. Thirteen FT/TFL1-like genes were isolated from petunia, with the five FT-like genes mainly expressed in leaves. The circadian rhythms of five FT-like genes and PhCO (petunia CONSTANS ortholog) were figured out. The expression of PhFT1 was contrary to that of PhFT2, PhFT3, PhFT4, and PhFT5. PhCO had a circadian clock different from Arabidopsis CO, but coincided with PhFT1; it decreased in daytime and accumulated at night. Two of the FT-like genes with differential circadian rhythm and higher expression levels, PhFT1 and PhFT4, were used to transform Arabidopsis. Eventually, overexpressing PhFT1 strongly delayed flowering, whereas overexpression of PhFT4 produced extremely early-flowering phenotype. Different from previous reports, PhTFL1a, PhTFL1b, and PhTFL1c were relatively highly expressed in roots. Taken together, this study demonstrates that petunia FT-like genes, like FT, are able to respond to photoperiod. The expression pattern of FT/TFL1 gene family in petunia contributes to a new insight into the functional evolution of this gene family.

FLOWERING LOCUS T (FT) is a key integrator of environmental signals and internal cues, and codes for florigen-like activity which regulates the transition from vegetative to reproductive growth in flowering plants. Unlike annual plants, perennial tree species undergo several years of vegetative growth prior to the transition to the reproductive stage, as characterised by the ability to form flower buds. Thereafter, trees in temperate regions typically display an annual growth cycle involving distinct vegetative growth, flowering and dormancy stages. In London plane (Platanus acerifolia Willd.), a FT-like gene has previously been identified. Here, we report the isolation of a novel FT orthologous gene, PaFTL, and investigate the functions of PaFT and PaFTL through the analysis of expression profiles and transgenic phenotypes. PaFT displayed the highest levels of expression during tree dormancy, and similarly elevated expression levels were seen under conditions of low temperature and short days (LT/SD). In contrast, PaFTL transcripts were up-regulated during the floral transition phase, the early stages of inflorescence development and throughout the main flowering period, whereas expression levels were low and variable during dormancy and in response to LT/SD treatments. Ectopic expression of 35s::PaFTL in tobacco produced a phenotype similar to that with PaFT, namely, advanced floral initiation. Overall, the results suggest that PaFT and PaFTL have both conserved and diverse functions in floral initiation, floral development and dormancy regulation.

In plants, most DREB1s/CBFs proteins with a single AP2/ERF domain participate in responses to abiotic stresses. Here, we cloned a DREB subfamily gene from Lilium longiflorum (designated as LlDREB1G). Sequence analyses revealed that it is composed of 219 amino acids, and contains an AP2/ERF domain, a putative nuclear localization signal (NLS), a DSAW/R and LWSY motif. LlDREB1G, which was localized to the nucleus, showed strongly induced expression under low temperature, and was also stimulated by mechanical agitation, heat shock, ABA and H2O2 stresses. Moreover, the LlDREB1G gene had an obvious circadian rhythm and organ specificity in expression. The LlDREB1G transcription factor was functional in the heterologous system (Arabidopsis). Transgenic Arabidopsis plants with higher transcription of LlDREB1G gene exhibited increased acquired thermotolerance and resistance to freezing and dehydration stresses. Besides, the transgenic seeds were insensitive to exogenous ABA treatment than those of the wild type (WT) in germination experiments. Compared with the WT seedlings, the seedlings of transgenic Arabidopsis overexpressing LlDREB1G showed strongly upregulated expression of stress-related genes after cold, drought or acclimation to heat shock treatments. Furthermore, overexpression of LlDREB1G increased the proline levels and survival rates, but decreased the electrolyte leakage and H2O2 accumulation in the transgenic plants after freezing, dehydration and high temperature treatments. These results imply that LlDREB1G gene may be involved in the responses to multiple abiotic stresses in an ABA-dependent and -independent manner. LlDREB1G gene, a DREB subfamily gene from Lilium longiflorum, may be involved in the responses to multiple abiotic stresses in an ABA-dependent and -independent manner.

Tall fescue is a widely used cool season turfgrass. In this study, we screened freezing tolerance of six tall fescue cultivars based on TEL50 (temperature causing 50% electrolyte leakage). We observed that freezing tolerant tall fescue cultivar exhibited lower EL, accumulated higher soluble sugar and proline, and showed higher CAT and SOD activities when compared with freezing intolerant cultivar. Based on bioinformatics analysis, 6 FaDREB1 unigenes were identified from publicly available transcriptomic data. Expression profiles showed that freezing tolerant cultivar only activated specific FaDREB1 unigenes, while freezing intolerant cultivar activated all six FaDREB1s during cold acclimation period. These findings indicated that cold stress modulated diverse physiological process in different tall fescue cultivars and activated various gene networks through regulating different subsets of FaDREB1s.

Roses are among the most economically important ornamental plants worldwide. But prickles on the stem and leaves cause difficulties for cultivation or inconveniences during harvest and transportation, thus are an undesirable horticultural character. However, little is known about the molecular mechanisms of prickle development. In this study, we sought to develop Rosa multiflora (in the family Rosaceae) as a model plant to study prickle formation. The morphology, structure, and ontogeny of prickles were characterized, and transcriptome analysis of prickly and prickleless R. multiflora genotypes was performed. Morphological observation and microscopic analyses revealed that prickles of R. multiflora were non-glandular prickles (NGPs) and their maturation went through five developmental stages, which was accompanied by the accumulation of secondary metabolites such as lignin and anthocyanins. Comparative transcriptome analysis identified key pathways and hub genes potentially involved in prickle formation. Interestingly, among the differentially expressed genes (DEGs), several notable development and secondary metabolism-related transcription factors (TFs) including NAC, TCP, MYB, homeobox, and WRKY were up-regulated in prickly internodes. KEGG enrichment analysis indicated that DEGs were enriched in the pathways related to biosynthesis of secondary metabolites, flavonoids, and phenylpropanoids in the prickly R. multiflora. Our study provides novel insights into the molecular network underlying the regulation of prickle morphogenesis in R. multiflora, and the identified candidates might be applied to the genetic improvement of roses.

Marigold (Tagetes erecta), as one member of Asteraceae family, bears a typical capitulum with two morphologically distinct florets. The SEPALLATA genes are involved in regulating the floral meristem determinacy, organ identity, fruit maturation, seed formation, and plant architecture. Here, five SEP-like genes were cloned and identified from marigold. Sequence alignment and phylogenetic analysis demonstrated that TeSEP3-1, TeSEP3-2, and TeSEP3-3 proteins were grouped into SEP3 clade, and TeSEP1 and TeSEP4 proteins were clustered into SEP1/2/4 clade. Quantitative real-time PCR analysis revealed that TeSEP1 and TeSEP3-3 were broadly expressed in floral organs, and that TeSEP3-2 and TeSEP4 were mainly expressed in pappus and corollas, while TeSEP3-1 was mainly expressed in two inner whorls. Ectopic expression of TeSEP1, TeSEP3-2, TeSEP3-3, and TeSEP4 in arabidopsis and tobacco resulted in early flowering. However, overexpression of TeSEP3-1 in arabidopsis and tobacco caused no visible phenotypic changes. Notably, overexpression of TeSEP4 in tobacco decreased the number of petals and stamens. Overexpression of TeSEP1 in tobacco led to longer sepals and simpler inflorescence architecture. The comprehensive pairwise interaction analysis suggested that TeSEP proteins had a broad interaction with class A, C, D, E proteins to form dimers. The yeast three-hybrid analysis suggested that in ternary complexes, class B proteins interacted with TeSEP3 by forming heterodimer TePI-TeAP3-2. The regulatory network analysis of MADS-box genes in marigold further indicated that TeSEP proteins played a "glue" role in regulating floral organ development, implying functional conservation and divergence of MADS box genes in regulating two-type floret developments. This study provides an insight into the formation mechanism of floral organs of two-type florets, thus broadening our knowledge of the genetic basis of flower evolution.

The flowering-time gene FD encodes a bZIP transcription factor that interacts with FLOWERING LOCUS T (FT) to induce flowering in Arabidopsis. Previous research has identified two FT homologs of Platanus acerifolia, PaFT and PaFTL, which each have different expression patterns and are involved in diverse developmental processes. However, it is not known whether such FT/FD complexes participate in the flowering processes in P. acerifolia. Therefore, we isolated two closely related FD homologs, PaFDL1 and PaFDL2, and investigated their functions through the analysis of expression profiles, transgenic phenotypes, their interactions with different FT proteins, and potential cis-regulatory elements in their promoters. The PaFDL genes were found to display their maximal expression levels during the stage of floral transition, and subsequent expression patterns were also seen to be related to inflorescence developmental stage. In addition, both PaFDL1 and PaFDL2 were found to be subject to post-transcriptional alternative splicing, each gene producing two transcript forms. Transgenic tobacco overexpressing each of the four resulting transcript types displayed accelerated floral initiation and produced abnormal flowers. The results suggested that the complete PaFDL proteins may interact with different PaFT/PaFTL proteins in order to fulfill both conservative and diverse functions in floral initiation and floral development.

Background AGAMOUS (AG) subfamily genes regulate the floral organs initiation and development, fruit and seed development. At present, there has been insufficient study of the function of AG subfamily genes in Asteraceae. Marigold (Tagetes erecta) belongs to Asteraceae family whose unique inflorescence structure makes it an important research target for understanding floral organ development in plants. Results Four AG subfamily genes of marigold were isolated and phylogenetically grouped into class C (TeAG1 and TeAG2) and class D (TeAGL11–1 and TeAGL11–2) genes. Expression profile analysis demonstrated that these four genes were highly expressed in reproductive organs of marigold. Subcellular localization analysis suggested that all these four proteins were located in the nucleus. Protein-protein interactions analysis indicated that class C proteins had a wider interaction manner than class D proteins. Function analysis of ectopic expression in Arabidopsis thaliana revealed that TeAG1 displayed a C function specifying the stamen identity and carpel identity, and that TeAGL11–1 exhibited a D function regulating seed development and petal development. In addition, overexpression of both TeAG1 and TeAGL11–1 leaded to curling rosette leaf and early flowering in Arabidopsis thaliana. Conclusions This study provides an insight into molecular mechanism of AG subfamily genes in Asteraceae species and technical support for improvement of several floral traits.

The plant-specific TCP transcription factors are well-characterized in both monocots and dicots, which have been implicated in multiple aspects of plant biological processes such as leaf morphogenesis and senescence, lateral branching, flower development and hormone crosstalk. However, no systematic analysis of the petunia TCP gene family has been described. In this work, a total of 66 petunia TCP genes (32 PaTCP genes in P. axillaris and 34 PiTCP genes in P. inflata) were identified. Subsequently, a systematic analysis of 32 PaTCP genes was performed. The phylogenetic analysis combined with structural analysis clearly distinguished the 32 PaTCP proteins into two classes&mdash, class &Iota, and class Ⅱ. Class Ⅱ was further divided into two subclades, namely, the CIN-TCP subclade and the CYC/TB1 subclade. Plenty of cis-acting elements responsible for plant growth and development, phytohormone and/or stress responses were identified in the promoter of PaTCPs. Distinct spatial expression patterns were determined among PaTCP genes, suggesting that these genes may have diverse regulatory roles in plant growth development. Furthermore, differential temporal expression patterns were observed between the large- and small-flowered petunia lines for most PaTCP genes, suggesting that these genes are likely to be related to petal development and/or petal size in petunia. The spatiotemporal expression profiles and promoter analysis of PaTCPs indicated that these genes play important roles in petunia diverse developmental processes that may work via multiple hormone pathways. Moreover, three PaTCP-YFP fusion proteins were detected in nuclei through subcellular localization analysis. This is the first comprehensive analysis of the petunia TCP gene family on a genome-wide scale, which provides the basis for further functional characterization of this gene family in petunia.

Sweetgums (Liquidambar), members of the family Altingiaceae (Altingiales), have inflorescences and floral organs that are distinctive in structure compared with other angiosperms in which the roles of floral homeotic genes have been studied. To begin to understand the role of AGAMOUS (AG)-a floral homeotic gene that has a major role in stamen and carpel development-in development of the monosexual flowers of sweetgum, we used RNAi to reduce the expression of two members of the AG subfamily. Because AG suppression should induce floral sterility, RNAi might also provide a tool to mitigate the risks of invasiveness-and to reduce the production of its nuisance fruits or allergenic pollen-when sweetgum is used as an exotic shade or forest tree. We tested 33 independent transgenic events and non-transgenic controls during 10 years in the field. The RNAi-AG sweetgum trees maintained normal growth, phenology, and vivid fall coloration during the 10 years of study, but 8 insertion events had highly modified inflorescence and floral morphology. The modified flowers had anthers and carpels that were converted to flat leaf-like structures lacking pollen grains and ovules, respectively. The female inflorescences developed into dry papery structures that failed to produce seeds. These infructescences were smaller than control infructescences, and lost a greater percentage of biomass in a controlled decay assay. RNAi against AG genes was highly effective at impairing fertility and modifying reproductive development without significant vegetative effects in sweetgum and gave phenotypes distinct from, but similar to, that of AG loss of function in other angiosperms. USDA-ARS, Mechanisms for Creating Non-Invasive Nursery Crops Nursery and Floriculture Initiative [1230-21000-051-06S]; J Frank Schmidt Charitable Foundation; Tree Biosafety and Genomics Research Cooperative This work was primarily funded by the USDA-ARS, Mechanisms for Creating Non-Invasive Nursery Crops Nursery and Floriculture Initiative, project No 1230-21000-051-06S. Funding was also provided by the J Frank Schmidt Charitable Foundation and the Tree Biosafety and Genomics Research Cooperative (formerly the Tree Genetic Engineering Research Cooperative).

Weeping is a specific plant architecture with high ornamental value. Despite the considerable importance of the weeping habit to landscaping applications and knowledge of plant architecture biology, little is known regarding the underlying molecular mechanisms. In this study, growth and phytohormone content were analyzed among the progeny of different branch types in an F1 mapping population of Prunus mume with varying plant architecture. Bulked segregant RNA sequencing was conducted to compare differences among progeny at a transcriptional level. The weeping habit appears to be a complex process regulated by a series of metabolic pathways, with photosynthesis and flavonoid biosynthesis highly enriched in differentially expressed genes between weeping and upright progeny. Based on functional annotation and homologous analyses, we identified 30 candidate genes related to weeping that merit further analysis, including 10 genes related to IAA and GA3 biosynthesis, together with 6 genes related to secondary branch growth. The results of this study will facilitate further studies of the molecular mechanisms underlying the weeping habit in P. mume.

An SNP mutation in the miR172 target site of the DcAP2L gene is associated with a double flower phenotype in Dianthus chinensis., The double flower is a highly important breeding trait that affects the ornamental value in many flowering plants. To get a better understanding of the genetic mechanism of double flower formation in Dianthus chinensis, we have constructed a high-density genetic map using 140 F2 progenies derived from a cross between a single flower genotype and a double flower genotype. The linkage map was constructed using double-digest restriction site-associated DNA sequencing (ddRAD-seq) with 2353 single nucleotide polymorphisms (SNPs). Quantitative trait locus (QTL) mapping analysis was conducted for 12 horticultural traits, and major QTLs were identified for nine of the 12 traits. Among them, two major QTLs accounted for 20.7% and 78.1% of the total petal number variation, respectively. Bulked segregant RNA-seq (BSR-seq) was performed to search accurately for candidate genes associated with the double flower trait. Integrative analysis of QTL mapping and BSR-seq analysis using the reference genome of Dianthus caryophyllus suggested that an SNP mutation in the miR172 cleavage site of the A-class flower organ identity gene APETALA2 (DcAP2L) is responsible for double flower formation in Dianthus through regulating the expression of DcAG genes.

In Platanus acerifolia, the SEP3 homolog PlacSEP3 may possess multiple functions, including roles in floral organ identity, development, and vegetative development. This work aimed to characterize the function of PlacSEP3 promoter and analyzed whether PlacSEP3 promoter could be used for genetic engineering flowerless London plane cultivars. The PlacSEP3 promoter was isolated by TAIL-PCR and cis-acting regulatory elements were predicted using the PLACE database and PlantCARE database. Three 5′-deletions of PlacSEP3 promoter were fused to the GUS gene and transformed into tobacco to analyze the functional by Histochemical staining and fluorometric determination. The pPlacSEP3-3::Barnase and pPlacSEP3-3::Barnase-mic35S-Barstar vectors were constructed and transformed into tobacco to test whether pPlacSEP3 could be used for genetic engineering flowerless London plane cultivars. A 1491-bp sequence was isolated, this sequence contained multiple putative cis-regulatory elements related to flower, pollen, and embryo/endosperm development, light responsive. Histochemical staining and fluorometric determination showed that GUS was strongly expressed in reproductive organs and apical buds, and was slightly expressed in vegetative tissues, except the roots of pPlacSEP3-1::GUS and pPlacSEP3-3::GUS transgenic tobacco. However, GUS was not expressed in floral organs and was just slightly expressed in fruits of pPlacSEP3-2::GUS transgenic tobacco. Interestingly, transgenic tobacco of all three constructs showed GUS staining at the bud side of semi-lignified and old stems. The pPlacSEP3-3::Barnase-mic35S-Barstar transgenic tobacco showed various degrees of sterility and vegetative developmental defects. The pPlacSEP3 involved in flower initiation, development and some aspects of vegetative development, such as lateral bud initiation and development. The pPlacSEP3 had the potential application for genetic engineering flowerless cultivars.

Chloroplasts convert solar energy into biologically useful forms of energy by performing photosynthesis. Although light and particular genes are known to promote chloroplast development, little is known about the mechanisms that regulate the tissue-specificity and cell-specificity of chloroplast biogenesis. Thus, the mechanisms that determine whether non-photosynthetic plastids rather than chloroplasts develop in petals remain largely unexplored. Although heat stress is known to inhibit photosynthesis, we do not know whether heat stress affects chloroplast biogenesis. Here, we report that heat stress up-regulates the expression of chlorophyll biosynthesis-related genes and promotes chloroplasts biogenesis in petals overexpressing SOC1 (suppressor of overexpression of CO) and novel SOC1-like genes. We also found that these specific MADS-box transcription factors are present in most photosynthetic eukaryotes and that the expression of more than one homolog is observed in chloroplast-containing tissues. These findings not only provide novel insights into the tissue specificity of chloroplast biogenesis and a method for producing green petals but also are consistent with heat stress influencing chloroplast biogenesis in higher plants.

A rapid, prolific and reproducible protocol for in vitro shoot regeneration from mature cotyledons of Platanus acerifolia has been developed. The influences of different plant growth regulator (PGR) combinations and donor seedling ages on shoot regeneration were investigated. The results showed that the application of BA in conjunction with NAA was the most effective PGR combination for the induction of shoot regeneration. When cotyledon explants of 5-day-old seedlings were incubated on MS basal medium supplemented with 4.0 mg L−1 BA and 0.2 mg L−1 NAA, 67.6 ± 4.9% of the cotyledon segments produced adventitious shoots. These regenerated shoots were initially formed as stunted rosette cluster forms and were encouraged to elongate to produce distinct shoots by transfer onto MS medium containing 0.5 mg L−1 BA and 0.05 mg L−1 NAA; the resulting mean number of adventitious shoots per explant was 5.81 ± 0.36. The elongated shoots were readily induced to root (i.e. 89.3% of shoots) by incubation on ½-strength MS medium supplemented with 0.1 mg L−1 IBA. This is the first report of an efficient in vitro shoot regeneration protocol for P. acerifolia through direct organogenesis using cotyledon explants. Hence, this provides a more efficient basis for the Agrobacterium-mediated genetic transformation of Platanus than previously available.

Breeding flowerless and/or fruitless varieties are highly desirable for London plane tree because it can prevent pollen- and fruit-mediated environmental contamination. Floral tissue-specific cell ablation is an efficient method to create such sterile plants. Here we isolated and characterized APETALA3 (AP3)-like and PISTILLATA (PI)-like genes and the promoters of PaAP3 and PaPI, in London plane tree respectively. The promoter fragments were fused to GUS (β-glucuronidase) and BARNASE gene, respectively, and transformed into tobacco plants. In pPaAP3::GUS transgenic plants, the GUS activity could be detected in various organs, including leaves, stems and all floral organs. Furthermore, most tobacco plants transformed with pPaAP3::BARNASE were dead and the survivals showed abortion of inflorescence. In contrast, heterologous expression of pPaPI::GUS in tobacco plants led to specific GUS activity in the inner three whorls of flowers. Accordingly, tobacco plants transformed with pPaPI::BARNASE lack petal, stamen and pistil, with only sepal left. The results suggest that sterile lines of P. acerifolia may be obtained by genetic engineering with pPaPI::BARNASE construct, which might solve the problems of shedding fruit hairs and disseminative pollens, reducing air pollution and reducing the allergens that harmful to human health.

Flowers with more petals are of more ornamental value. It is well known that AGAMOUS (AG) is the core member of the C-class gene which plays an essential role in double flower formation and identification of stamens and carpels in Arabidopsis thaliana. We searched C-class genes in the genome of the carnation, and found two AG orthologs (DcaAGa, DcaAGb). Phylogenetic analysis showed that the two genes were closely related to the euAG subclade. Then we searched the genomes of other Caryophyllales plants (Beta vulgaris, Spinacia oleracea, Chenopodium quinoa) for C-class genes, and found that their C-class genes all belonged to the euAG subclade. Semi-quantitative PCR (sq-PCR) analysis indicated that the expression of DcaAG genes in the single flower phenotype was higher than that in the double flower phenotype. Quantitative real-time RT-PCR (qRT-PCR) analysis showed that the expressions of DcaAG genes in the flower bud were significantly different from those in the root, stem, and leaf between the single and double flower phenotype carnations, and that DcaAG genes were specifically expressed in the stamen and carpel of carnation. Moreover, the expression of other floral organ identity genes (AP1 and AP2, PI and AP3, SEP1 and SEP3 corresponding to the A-, B-, and E-class of genes, respectively) showed no significant difference in all floral organs between the single and double flower phenotype carnations, suggesting that C-class (DcaAG) genes might play an important role in the double flower phenotype in carnation. Petal loss or decrease, precocious flowering, silique shortening, and seed sterility were observed in 35S::DcaAGa and 35S::DcaAGb transgenic Arabidopsis plants. All these results show that DcaAG genes might affect the petal number negatively and have a specific function in stamen and carpel development in carnation.

Additional file 2: Table S2. Amino acid sequence alignment of C class proteins.

Additional file 11: Table S8. Raw data of CT value in qRT-PCR for expression levels of AP1, AP3, PI, AG, and STK in flowers from 35S:TeAGL11–1 transgenic lines and wild-type Arabidopsis.

Additional file 6: Fig. S3. Expression of TeAG1, TeAG2, TeAGL11–1 and TeAGL11–2 in seedlings of T1 transgenic lines by semi-quantitative RT-PCR. (a-1) 35S:TeAG1 transgenic lines. (b-1) 35S:TeAG2 transgenic lines. (c-1) 35S:TeAG11–1 transgenic lines. (d-1) 35S:TeAG11–2 transgenic lines. WT: wild type line; SL: strong phenotypic line; WL: weak phenotypic line; L: transgenic line. (a-2, b-2, c-2, d-2), the constitutive gene is Arabidopsis keeping-house gene AtEF1α. (DOCX 654 kb)

Additional file 4: Fig. S1. Expression levels of TeAG1, TeAG2, TeAGL11–1 and TeAGL11–2 in different tissues and organs. Rt: root; Sm: stems; Le: leaf; FB1-FB4: flower buds were 0-1 mm, 2-3 mm, 4–5 mm and 6-7 mm in diameter, respectively; Re: receptacle; Br: bract; RS: sepal of ray floret; RP: petal of ray floret; RPi: pistil of ray floret; Se: sepal of disk floret; Pe: petal of disk floret; St: stamen of disk floret; Pi: pistil of disk floret; Ov: ovary.

Additional file 7: Table S4. Statistics for seed setting rate between control and Sl-TeAG1 transgenic lines.

Additional file 3: Table S3. Amino acid sequence alignment of D class proteins.

Additional file 9: Table S6. Raw data of CT value in qRT-PCR for expression levels of TeAG1, TeAG2, TeAGL11–1, and TeAGL11–2 in different tissues and organs of marigold.

Flower senescence is classified into ethylene-dependent and ethylene-independent manners and determines the flower longevity which is valuable for ornamental plants. However, the manner of petal senescence in tulip is still less defined. In this study, we characterized the physiological indexes in the process of petal senescence, as well as metabolic and ethylene responses in tulip cultivar 'American Dream', and further identified the role of ethylene biosynthesis genes TgACS by transgenic and transient assays. Primary metabolites profiling revealed that sugars, amino acids and organic acids preferentially accumulated in senescent petals. Additionally, senescence-associated genes were identified and significantly up-regulated, coupled with increased ROS contents, rapid water loss and accelerated cell membrane breakdown. Moreover, ethylene production was stimulated as evidenced by increasing in ACS activity and ethylene biosynthesis-related genes expression. Exogenous treatment of cutting flowers with 1-MCP or ethephon resulted in delayed or enhanced petal senescence, respectively. Transient down-regulation of TgACS by VIGS assay in tulip petals delayed senescence, while over-expressed TgACS1 in tobacco promoted leaf senescence. Taken together, this study provides evidences to certify ethylene roles and TgACS functions during flower senescence in tulip.

Lycopene e-cyclases (LCYEs) are key enzymes in carotenoid biosynthesis converting red lycopene to downstream lutein. The flowers of marigold (Tagetes erecta) have been superior sources to supply lutein. However, the transcriptional regulatory mechanisms of LCYe in lutein synthesis are still unclear in marigold. In this work, the expression pattern of the TeLCYe gene in marigold indicated that TeLCYe mainly expressed in floral organs. To gain a better understanding of the expression and regulatory mechanism of TeLCYe gene, the TeLCYe promoter was isolated, sequenced, and analyzed through bioinformatics tools. The results suggested that the sequence of TeLCYe promoter contained various putative cis-acting elements responsive to exogenous and endogenous factors. The full-length TeLCYe promoter and three 5′-deletion fragments were fused to the GUS reporter gene and transferred into tobacco to test the promoter activities. A strong GUS activity was observed in stems of seedlings, leaves of seedlings, middle stems, top leaves, petals, stamens, and stigmas in transgenic tobacco containing full-length TeLCYe promoter LP0-2086. Deletion of − 910 to − 429 bp 5′ to ATG significantly increased the GUS activity in chloroplast-rich tissues and floral organs, while deletion occurring from 1170 to 910 bp upstream ATG decreased the TeLCYe promoter strength in stems of seedlings, leaves of seedlings, top leaves and sepals. Functional characterization of the full-length TeLCYe promoter and its’ deletion fragments in stable transgenic tobacco indicated that the LP0-2086 contains some specific cis-acting elements, which might result in the high-level expression of in floral organs, and LP2-910 might contain some specific cis-acting elements which improved GUS activities in vegetable tissues.

LEAFY (LFY) and UNUSUAL FLORAL ORGANS (UFO) homologous genes have been reported to play key roles in promoting the initiation of floral meristems in raceme- and cyme-type plants. Asteraceae, a large family of plants with more than 23,000 species, has a unique head-like inflorescence termed capitulum. Here, we report a floral defective plant of the garden cosmos named green head (gh), which shows homogeneous inflorescence, indistinguishable inflorescence periphery and center, and the replacement of flower meristems by indeterminate inflorescence meristems, coupled with iterative production of bract-like organs and higher order of inflorescences. A comparison of the LFY- and UFO-like genes (CbFLY and CbUFO) isolated from both the wild-type and gh cosmos revealed that CbUFO may play an important role in inflorescence differentiation into different structures and promotion of flower initiation, and the reduced expression of CbUFO in the gh cosmos could be associated with the phenotypes of the flower-defective plants. Further expression analysis indicated that CbUFO may promote the conversion of inflorescence meristem into floral meristem in early ray flower formation, but does not play a role in its later growth period.

LEAFY (LFY) and UNUSUAL FLORAL ORGANS (UFO) homologous genes have been reported to play key roles in promoting the initiation of floral meristems in raceme- and cyme-type plants. Asteraceae, a large family of plants with more than 23,000 species, has a unique head-like inflorescence termed capitulum. Here, we report a floral defective plant of the garden cosmos named green head (gh), which shows homogeneous inflorescence, indistinguishable inflorescence periphery and center, and the replacement of flower meristems by indeterminate inflorescence meristems, coupled with iterative production of bract-like organs and higher order of inflorescences. A comparison of the LFY- and UFO-like genes (CbFLY and CbUFO) isolated from both the wild-type and gh cosmos revealed that CbUFO may play an important role in inflorescence differentiation into different structures and promotion of flower initiation, and the reduced expression of CbUFO in the gh cosmos could be associated with the phenotypes of the flower-defective plants. Further expression analysis indicated that CbUFO may promote the conversion of inflorescence meristem into floral meristem in early ray flower formation, but does not play a role in its later growth period.

Tagetes erecta is an annual multifunctional plant which can be cultivated under a broad range of climatic conditions. Polyploidization and interspecific hybridization are applied to facilitate breeding cultivars of T. erecta with improved ornamental qualities. Colchicine treatment to the germinating seeds was proved to be a useful tool for chromosome doubling of the male sterile two-type line ‘M525AB’, with the resulting frequency of polyploid seedlings ranging from 88.89% (following 0.05% w/v colchicine applied for a 3–6 h exposure period) to a maximum of 100.00% (following 0.1% for 3–6 h, or 0.2% for 3 h). Morphological observation, stomatal size and density analysis, flow cytometric analysis and chromosome counting were conducted to identify the tetraploid plants. Distinctive morphological changes were observed in a notable proportion of polyploid plants. The colchicine-treated polyploid T. erecta plants showed dwarfed and more robust growth, thicker, larger and greener leaves, larger inflorescences and florets. The mutant plants identified through morphological observation all aligned as polyploid plants, thus morphological observation could be an effective method for the detection of polyploidy. The polyploid plants had significant larger stomata size over the abaxial leaf surface, whereas the density of stomata distribution was remarkably reduced. The survival rate of tetraploid cuttings (i.e. 38%) was greatly reduced compared to that of diploid plants. The fertility of tetraploid plants was also decreased, as shown by cross-pollination yields. Interspecific hybridizations between colchicine-induced tetraploid plants of a male sterile T. erecta line and the naturally tetraploid fully fertile Tagetes patula species resulted in hybrid progeny. Most of these hybrids displayed the dwarfed growth stature and compact, larger-flower morphology which is the typical ideotype of herbaceous flowers. Thus, polyploidization may be employed effectively as a means to facilitate interspecific hybridization, thereby contributing significantly to the improvement of quantitative traits of Tagetes spp.

The flower architectural trait of doubleness is a highly important breeding characteristic in the ornamental species petunia (Petunia hybrida). Here, we described the development of a near-isogenic line of a petunia population that segregated for single and double flower types. Sequence-related amplified polymorphism (SRAP) and simple sequence repeat (SSR) techniques, combined with bulked segregant analysis (BSA) were used to develop molecular markers linked to this floral trait. From a survey of 170 SRAP and 38 SSR primer combinations, one SRAP marker and one SSR marker were found to be linked to the target trait, each leading to the amplification of a specific band in double flower petunia. The two markers were verified in a population of 1065 plants, including 478 single flower and 587 double flower petunia individuals, and were shown to be tightly linked to the double flower locus with no recombination events observed. Hence, these markers have the potential for application in the positional analysis of the double flower locus and, ultimately, may contribute towards determining the genetic basis of flower type.

Mei (Prunus mume Sieb. et Zucc.) is an important ornamental plant in China, but low temperature severely restricts its application in North China. The C-repeat binding factor/Dehydration responsive element binding factors (CBF/DREB1) pathway has important roles in plant responses to cold stress. However, knowledge on the function of CBF/DREB1 genes from Mei and its regulatory roles in freezing and other abiotic stresses is limited. Three PmhCBF genes, designated as PmhCBFa, PmhCBFb and PmhCBFc respectively, were isolated and characterized from Prunus mume ‘Huahudie’. Gene expression analysis revealed that PmhCBFa and PmhCBFb were more induced than PmhCBFc by cold stress, and other stress treatments including H2O2, Mannitol, NaCl and Abscisic Acid also induced the expressions of PmhCBFa/b/c at a relatively lower level. Compared to PmhCBFa and PmhCBFb, PmhCBFc appeared to be more triggered by H2O2 and showed broad regulation potential in all the abiotic stresses. Overexpression of PmhCBFc in Arabidopsis improved freezing and oxidative stress tolerance, and reduced the inhibition of PSII, but delayed development and flowering. The superoxide dismutase (SOD, EC 1.11.1.6) and peroxidase (POD, EC 1.11.1.7) activities were also elevated in the transgenic Arabidopsis plants during freezing-thawing cycles. Our results suggested that PmhCBFc responded to reactive oxygen species (ROS) and functioned in freezing tolerance by enhancing the capacities for ROS scavenging.

Four SPL3/4/5 orthologous genes from Platanus acerifolia were identified, and their structure, expression patterns, role in controlling phase change and ability of inducing early flowering in Arabidopsis were analyzed. Plant flowering behavior is controlled by multiple pathways. The Arabidopsis thaliana SQUAMOSA PROMOTER BINDING PROTEIN-LIKE3/4/5 (AtSPL3/4/5) genes are involved in regulating vegetative phase change. Recent studies have shown that AtSPL3/4/5 genes are directly involved in inducing expression of three flowering integrator genes namely, LEAFY (LFY), FRUITFULL (FUL) and APETALA1 (AP1), thereby triggering flowering. This regulatory integration reveals a commonality of the flowering response with various other plant developmental pathways. We have isolated four SPL3/4/5 orthologous genes (PaSPL3a/b/c/d) from a basal eudicot London plane tree (Platanus acerifolia Wild), a woody species with a long juvenile phase. We investigated the role of these Platanus genes in flowering and detected expression levels between trees in vegetative and reproductive phase. Real-time PCR analysis demonstrated diverse expression patterns for the four PaSPL3 genes throughout the various P. acerifolia tissues, but all four were highly expressed in developing flower bud tissues. Expression of PaSPL3a/b/c in leaves changed according to the season and was regulated by temperature. Furthermore, three of the orthologous were found to induce an early flowering phenotype when overexpressed in Arabidopsis. The findings of this study indicate that PaSPL3 genes may participate in early stage inflorescence development in London Plane. However, there appears to be some divergence in the function of these genes in the control of phase change in London plane as compared to that within Arabidopsis.

Additional file 3. Primers used in this study for cloning PhLSH CDS, qRT-PCR and transgene line RT-PCR.

Summary Flavonoids play critical roles in plant responses to various stresses. Few studies have been reported on what the mechanism of activating flavonoid biosynthesis in plant responses to wounding and oxidation is. In this study, flavonoid metabolites and many MYB transcript factors from Rosa rugosa were verified to be induced by wounding and oxidation. RrMYB5 and RrMYB10, which belong to PA1‐ and TT2‐type MYB TFs, respectively, showed extremely high induction. Overexpression of RrMYB5 and RrMYB10 resulted in an increased accumulation of proanthocyanidins in R. rugosa and tobacco by promoting the expression of flavonoid structural genes. Transcriptomic analysis of the transgenic plants showed that most genes, involved in wounding and oxidation response and ABA signalling modulation, were up‐regulated by the overexpression of RrMYB10, which was very much similar to that observed in RrANR and RrDFR overexpression transgenics. RrMYB5 and RrMYB10 physically interacted and mutually activated each other's expressions. They solely or synergistically activated the different sets of flavonoid pathway genes in a bHLH TF EGL3‐independent manner. Eventually, the accumulation of proanthocyanidins enhanced plant tolerance to wounding and oxidative stresses. Therefore, RrMYB5 and RrMYB10 regulated flavonoid synthesis in feedback loop responding to wounding and oxidation in R. rugosa. Our study provides new insights into the regulatory mechanisms of flavonoid biosynthesis by MYB TFs and their essential physiological functions in plant responses to wounding and oxidative stresses.

Four R3 MYB genes were cloned and identified from Platanus acerifolia and analysed according to endogenous gene expression profiles, protein–protein interaction patterns, phenotypic effects and related gene expression profiles in transgenic Arabidopsis, suggesting that London plane R3 MYB genes inhibit trichome formation in Arabidopsis. The CPC-like MYB transcription factors including CAPRICE (CPC), TRIPTYCHON (TRY), ENHANCER OF TRY AND CPC 1, 2 and 3 (ETC1, ETC2 and ETC3), TRICHOMELESS1 (TCL1) and TRICHOMELESS2(TCL2) play important roles in controlling trichome patterning in Arabidopsis. In this study, four sequences homologous with the Arabidopsis CPC family were identified from London plane and named PaTRY, PaCPC-like1, PaCPC-like2 and PaCPC-like3. Over-expression of PaTRY, PaCPC-like1, PaCPC-like2 and PaCPC-like3 in Arabidopsis resulted in glabrous phenotypes. In addition, expression of endogenous GL2, GL1, MYB23, TTG2 and a set of R3 MYB-encoding genes was markedly reduced. Furthermore, the protein products of PaTRY, PaCPC-like1, PaCPC-like2 and PaCPC-like3 were shown to interact with PaGL3 in yeast two-hybrid assays. Together, these results likely suggest that the mechanisms of trichome regulation in London plane have similarities with those in Arabidopsis.

The function of euAP1 and euFUL in AP1/FUL lineage have been well characterized in core eudicots, and they play common and distinct roles in plant development. However, the evolution and function of FUL-like genes is poorly understood in basal eudicots. In this study, we identified three FUL-like genes PlacFL1/2/3 from London plane (Platanus acerifolia). Sequence alignment and phylogenetic analysis indicated that PlacFL1/2/3 are AP1/FUL orthologs and encoded proteins containing FUL motif and paleoAP1 motif. Quantitative real-time PCR (qRT-PCR) analysis showed that PlacFL1/2/3 were expressed in both vegetative and reproductive tissues, but with distinct spatiotemporal patterns. In contrast to PlacFL1 and PlacFL3, PlacFL2 exhibited higher expression levels and broader expression regions, and that the expression of PlacFL2 gene showed a decreasing and increasing tendency in subpetiolar buds during dormancy induction and breaking, respectively. Overexpression of PlacFLs in Arabidopsis and PlacFL3 in tobacco resulted in early flowering, as well as early termination of inflorescence meristems for transgenic Arabidopsis plants. The expression changes of flowering time and flower meristem identity genes in transgenic Arabidopsis lines with different PlacFLs suggested that PlacFL2 and PlacFL3 may regulate different downstream genes to perform divergent functions. Yeast two-hybrid analysis indicated that PlacFLs interacted strongly with PlacSEP proteins, and PlacFL3 instead of PlacFL1 and PlacFL2 could also form a homodimer and interact with D-class proteins. Our results suggest that PlacFLs may play conserved functions in regulating flowering and flower development, and PlacFL2 might also be involved in dormancy regulation. The research helps us to understand the functional evolution of FUL-like genes in basal eudicots, especially in perennial woody species.

International audience; Dianthus is a large genus containing many species with high ornamental economic value. Extensive breeding strategies permitted an exploration of an improvement in the quality of cultivated carnation, particularly in flowers. However, little is known on the molecular mechanisms of flower development in carnation. Here, we report the identification and description of MADS-box genes in carnation (DcaMADS) with a focus on those involved in flower development and organ identity determination. In this study, 39 MADS-box genes were identified from the carnation genome and transcriptome by the phylogenetic analysis. These genes were categorized into four subgroups (30 MIKCc, two MIKC*, two M alpha, and five M gamma). The MADS-box domain, gene structure, and conserved motif compositions of the carnation MADS genes were analysed. Meanwhile, the expression of DcaMADS genes were significantly different in stems, leaves, and flower buds. Further studies were carried out for exploring the expression of DcaMADS genes in individual flower organs, and some crucial DcaMADS genes correlated with their putative function were validated. Finally, a new expression pattern of DcaMADS genes in flower organs of carnation was provided: sepal (three class E genes and two class A genes), petal (two class B genes, two class E genes, and one SHORT VEGETATIVE PHASE (SVP)), stamen (two class B genes, two class E genes, and two class C), styles (two class E genes and two class C), and ovary (two class E genes, two class C, one AGAMOUS-LIKE 6 (AGL6), one SEEDSTICK (STK), one B sister, one SVP, and one Ma). This result proposes a model in floral organ identity of carnation and it may be helpful to further explore the molecular mechanism of flower organ identity in carnation.

Background SQUAMOSA PROMOTER BINDING PROTEIN (SBP)-box genes encode a family of plant-specific transcription factors (TFs) that play important roles in many growth and development processes including phase transition, leaf initiation, shoot and inflorescence branching, fruit development and ripening etc. The SBP-box gene family has been identified and characterized in many species, but has not been well studied in Petunia, an important ornamental genus. Results We identified 21 putative SPL genes of Petunia axillaris and P. inflata from the reference genome of P. axillaris N and P. inflata S6, respectively, which were supported by the transcriptome data. For further confirmation, all the 21 genes were also cloned from P. hybrida line W115 (Mitchel diploid). Phylogenetic analysis based on the highly conserved SBP domains arranged PhSPLs in eight groups, analogous to those from Arabidopsis and tomato. Furthermore, the Petunia SPL genes had similar exon-intron structure and the deduced proteins contained very similar conserved motifs within the same subgroup. Out of 21 PhSPL genes, fourteen were predicted to be potential targets of PhmiR156/157, and the putative miR156/157 response elements (MREs) were located in the coding region of group IV, V, VII and VIII genes, but in the 3’-UTR regions of group VI genes. SPL genes were also identified from another two wild Petunia species, P. integrifolia and P. exserta, based on their transcriptome databases to investigate the origin of PhSPLs. Phylogenetic analysis and multiple alignments of the coding sequences of PhSPLs and their orthologs from wild species indicated that PhSPLs were originated mainly from P. axillaris. qRT-PCR analysis demonstrated differential spatiotemperal expression patterns of PhSPL genes in petunia and many were expressed predominantly in the axillary buds and/or inflorescences. In addition, overexpression of PhSPL9a and PhSPL9b in Arabidopsis suggested that these genes play a conserved role in promoting the vegetative-to-reproductive phase transition. Conclusion Petunia genome contains at least 21 SPL genes, and most of the genes are expressed in different tissues. The PhSPL genes may play conserved and diverse roles in plant growth and development, including flowering regulation, leaf initiation, axillary bud and inflorescence development. This work provides a comprehensive understanding of the SBP-box gene family in Petunia and lays a significant foundation for future studies on the function and evolution of SPL genes in petunia. Electronic supplementary material The online version of this article (10.1186/s12864-018-4537-9) contains supplementary material, which is available to authorized users.

Small heat shock proteins (sHSPs) have been shown to be involved in stress tolerance. However, their functions in Prunus mume under heat treatment are poorly characterized. To improve our understanding of sHSPs, we cloned a sHSP gene, PmHSP17.9, from P. mume. Sequence alignment and phylogenetic analysis indicated that PmHSP17.9 was a member of plant cytosolic class III sHSPs. Besides heat stress, PmHSP17.9 was also upregulated by salt, dehydration, oxidative stresses and ABA treatment. Leaves of transgenic Arabidopsis thaliana that ectopically express PmHSP17.9 accumulated less O2 − and H2O2 compared with wild type (WT) after 42 °C treatment for 6 h. Over-expression of PmHSP17.9 in transgenic Arabidopsis enhanced seedling thermotolerance by decreased relative electrolyte leakage and MDA content under heat stress treatment when compared to WT plants. In addition, the induced expression of HSP101, HSFA2, and delta 1-pyrroline-5-carboxylate synthase (P5CS) under heat stress was more pronounced in transgenic plants than in WT plants. These results support the positive role of PmHSP17.9 in response to heat stress treatment.

Cloning and functional analysis of CcBBM gene. Baby boom (BBM), as a transcription factor of AP2/ERF family, has great potence in promoting somatic embryogenesis. In this study, a BBM gene from somatic embryos of camphor tree (Cinnamomum camphora L.), named CcBBM, was cloned and characterized. A partial fragment of 560 bp of CcBBM was amplified by degenerate PCR based on the two conserved AP2 domains of other annotated BBMs. Full-length cDNA of CcBBM was obtained by a cloning strategy combing degenerate PCR and RACE technique. The open-reading frame of CcBBM is 2169 bp in length, which encodes 722 amino acids. A comparison of the deduced amino acid sequence of CcBBM with BBM from other species showed high similarities, ranging from 51 to 80 %. The mRNA levels of CcBBM in different tissues were detected by semi-quantitative reverse transcription PCR (RT-PCR). CcBBM was expressed in most tissues, with highest expression level in somatic embryos, followed by roots, young fruits and flowers. Ectopic expression of CcBBM gene in Arabidopsis exhibited a variety of deformed and variegated leaves and short roots. No spontaneous somatic embryogenesis was observed in 35S::CcBBM transgenic plants. These results will help us understand the role of CcBBM in somatic embryogenesis in camphor tree.

Rose (Rosa hybrida) is one of the most economically important ornamentals worldwide. Powdery mildew (Podosphaera pannosa) is a major disease in cut and potted roses. In dicots such as arabidopsis (Arabidopsis thaliana), pea (Pisum sativum), and tomato (Solanum lycopersicum), loss-of-function mutations in mildew resistance locus o (Mlo) genes confer high levels of broad-spectrum resistance to powdery mildew. Here, we present spatiotemporal expression patterns of four Mlo genes from R. hybrida based on real-time fluorescence quantitative polymerase chain reaction (qPCR). Phylogenetically closely related R. hybrida mildew resistance locus o (RhMLO) genes showed similar or overlapping tissue specificity and analogous responsiveness to external stimuli. RhMLO1 and RhMLO2 transcriptional levels were upregulated more than 2-fold by external stimuli, especially by inoculation with powdery mildew fungus P. pannosa at early time points. This phenomenon was not found for RhMLO3 or RhMLO4. The results indicated that RhMLO1 and RhMLO2 might play important roles in rose–powdery mildew pathogen interactions. Our findings may provide useful information for the study of mechanisms of powdery mildew susceptibility in rose.

Powdery mildew is one of the most important crop diseases worldwide. Genetic analysis has revealed that mutant alleles of the Mlo gene cause broad-spectrum resistance against pathogens found in cereal. In this study, the possibility of inducing powdery mildew (Podosphaera pannosa) resistance via transgenic technology in the rose was examined. The transgenic lines were confirmed for integration and copy number of the transgenes based on PCR and Southern blots. A clear correlation was found between resistance and the accumulation of silenced RhMLO1, based on real-time fluorescent quantitative PCR and resistance analysis. Compared with the control plants, transgenic roses displayed higher resistance levels. Furthermore, the results indicated that RhMLO1 has a negative role in the rose–powdery mildew pathogen interaction. This demonstrates the potentially viable strategy of rendering the Mlo homologs partially non-functional via a transgenic approach or mutagenesis in order to counter powdery mildew in Rosa. To the best of our knowledge, this study is one of the first attempts at using a Mlo-based resistance strategy to combat powdery mildew in Rosa.

The SANT/MYB transcription factors play a critical role in regulating the development of plants. Recently, various SANT/MYB transcription factors have been reported in some plant species, while none in petunia. In this research, a gene containing the SANT/MYB region, designated as PhRL , was firstly identified in petunia flower buds cDNA library. The open-reading frame of PhRL was 288 bp, encoding a putative 10.97 kDa protein of 95 amino acids. Sequence alignment exhibited that PhRL contained a high conserved SANT/MYB domain. Phylogenetic comparison showed that PhRL was clustered together with AtRL5 and AtRL6 of Arabidopsis . Different from the reported SANT/MYB transcription factors, PhRL was preponderantly expressed in the anther and then in the pistil, with little/no detection in sepal, petal, stem, root or leaf. RNA interference (RNAi) of the PhRL expression in petunia showed significant decrease in pollen viability, and enlargement in size of sex organs. These results indicated that PhRL was a fertility-related gene, which might be involved in both the anther and pistil development of petunia. Studies on PhRL may further our understanding of the molecular mechanisms of sex organs development in petunia.