Fig 1.
Stages of olive flower development corresponding to the sampling dates.
The sampling period includes the differentiation of flower clusters from inflorescence buds and the subsequent stages until the anther dehiscence of the developed flowers. Pictures of entire flower clusters (below) and single flower buds (above) are shown. Stages 1–6: developing flowers at pre-anthesis (26 to 3 days before anthesis). Stages 7–10: flowers at anthesis (0 to 3 days after anthesis). Stages b1-b3 represent the inflorescence buds collected at 42, 36 and 32 days before anthesis and were only used for the RT-qPCR experiments on the candidate homeotic genes.
Table 1.
An overview of the raw data from the 454-FLX Titanium sequencing.
Table 2.
An overview of the assemblies that putatively referred to flower development (FD), ovary abortion (OA) and pollen-pistil interactions (PPI).
Fig 2.
Clustering of the differentially expressed TCs.
(A) Hierarchical Clustering Analysis (HCA). The different shades of green and red represent the down- and up-regulation levels, respectively. (B) Principal Component Analysis (PCA). The percentage of variance explained by each component is shown within brackets. (C) Clustering of the co-expressed transcripts. L1-L6 = Leccino at pre-anthesis, DA1-DA6 = Dolce Agogia at pre-anthesis, L7-L10 = Leccino at anthesis, and F7-F10 = Frantoio at anthesis.
Fig 3.
The MapMan overview maps related to metabolism, regulatory network, cellular response and transcription show differences in the transcript levels between the Leccino flowers at pre-anthesis (stages 1–6) and anthesis (stages 7–10).
Fig 4.
GO terms for biological processes.
Distribution of the differentially expressed transcripts related to A—flower development, B—ovary abortion, and C—pollen-pistil interactions in the main GO categories for “biological process”, “molecular function” and “cellular component”.
Fig 5.
Expression of floral organ identity genes.
The genes involved in floral organ identity, according to the ABC model, are expressed in the olive flowers. (A) Schematic representation of the classical ABC model for the development of flower organs. Class A genes (red) specify sepals, class A and B (yellow) genes specify petals, class B and C (blue) genes specify stamens, class C genes specify carpels, class D genes (grey) are specifically expressed in ovules, and class E genes (green) have partially redundant functions in floral organ identity determination. (B) The un-rooted NJ tree of key determinants of floral organ identity emphasizes the relationships between the different ABCDE-class genes. The five main protein groups that were identified are indicated with different colours: SEPALLATA (SEP, green), APETALA1 (AP1, red), AGAMOUS (AG, blue), PISTILLATA (PI, yellow) and APETALA3 (AP3, brown). The green circles, red rectangles and blue triangles represent the proteins from Olea europaea, Antirrhinum majus and Arabidopsis thaliana, respectively. (C, D) Relative mRNA levels of 12 MADS-box genes in single flower parts (Ca/Co: calix and corolla, anther, pistil) of flowers at S4 stage (C) or whole inflorescence buds (b1-b3) and differentiated flowers (stages 1–3) at progressive developmental stages (D) of Leccino. OeAP1: unigene03554; OeAP2: unigene00847; OeFUL: FRUITFULL, unigene01990; OeAP3: unigene04520; OePI: unigene04760; OeAG: unigene03555; OeSHP1: SHATTERPROOF, unigene03887; OeSTK: SEEDSTICK, unigene07062; OeSEP2.1: unigene04526; OeSEP2.2: unigene02210; OeSEP3: unigene03885; and OeSEP4: unigene04898. The data were obtained using RT-qPCR. The values are the means of three biological replicates and the SD is indicated.
Table 3.
Genes involved in ABC model flower.
Table 4.
Genes involved in ovary abortion.
Fig 6.
Expression of ovary abortion genes.
Relative mRNA levels of the genes potentially related to ovary abortion, as determined by RT-qPCR. Flowers at stages 6 and 9 were analysed. OeGBSSI: Granule bound starch synthase I, unigene00185; OeSuSy: sucrose synthase, unigene02089; OeBAM1: β-amylase1, unigene01162; OeInv-CW: cell-wall invertase, unigene02494; OeInv-V: soluble acid invertase, unigene01665; OeADC: Arginine decarboxylase, unigene00039; OeNLP1: N-carbamoylputrescine amidase, unigene03141; OeSPDS: Spermidine synthase, unigene01406; OeSPMS: Spermine synthase, unigene06009; OeMYBPA1: unigene03795; and OeMYBC2: unigene04022. The transcripts were detected in pistils and anthers of perfect flowers, anthers of staminate flowers and entire undeveloped flowers. The values are means of three biological replicates and the SD is indicated.
Table 5.
Genes involved in pollen-pistil interaction.
Fig 7.
Expression of pollen-pistil interaction genes.
Relative mRNA levels, determined by RT-qPCR, of the genes that may be involved in pollen-pistil interactions, (A) in anthers and pistils at anthesis (stages 7–10), (B) in anthers before anthesis (stage 5) and at anthesis (stages 7–8), from flowers of Leccino and Frantoio. Transcripts analysed include: Lipid transfer proteins (OeLTP1: unigene05516; OeLTP2: unigene08514; OeLTP3: unigene07617; OeLTP4: unigene07897, unigene07066); pollen-specific gene (OeLAT52: unigene06374); pollen-specific gene (OeNTP: unigene03925); Pectin methylesterase (OePME1: unigene01185; OePME2: unigene01763). Oleosin (OeOLE: unigene08062); and β-1,3-glucanase (OeβGLU: unigene00872). The values are the means of three biological replicates; the SD is indicated.