Variation in shape and size of many floral organs is related to pollinators. Evolution of such organs is driven by duplication and modification of MADS-box and MYB transcription factors. We applied a combination of micro-morphological (SEM and micro 3D-CT scanning) and molecular techniques (transcriptome and RT-PCR analysis) to understand the evolution and development of the callus, stelidia and mentum, three highly specialized floral structures of orchids involved in pollination. Early stage and mature tissues were collected from flowers of the bee-pollinated Phalaenopsis equestris and Phalaenopsis pulcherrima, two species that differ in floral morphology: P. equestris has a large callus but short stelidia and no mentum, whereas P. pulcherrima has a small callus, but long stelidia and a pronounced mentum. Our results show the stelidia develop from early primordial stages, whereas the callus and mentum develop later. In combination, the micro 3D-CT scan analysis and gene expression analyses show that the callus is of mixed petaloid-staminodial origin, the stelidia of staminodial origin, and the mentum of mixed sepaloid-petaloid-staminodial origin. SEP clade 1 copies are expressed in the larger callus of P. equestris, whereas AP3 clade 1 and AGL6 clade 1 copies are expressed in the pronounced mentum and long stelidia of P. pulcherrima. AP3 clade 4, PI-, AGL6 clade 2 and PCF clade 1 copies might have a balancing role in callus and gynostemium development. There appears to be a trade-off between DIV clade 2 expression with SEP clade 1 expression in the callus, on the one hand, and with AP3 clade 1 and AGL6 clade 1 expression in the stelidia and mentum on the other. We detected differential growth and expression of MADS box AP3/PI-like, AGL6-like and SEP-like, and MYB DIV-like gene copies in the callus, stelidia and mentum of two species of Phalaenopsis, of which these floral structures are very differently shaped and sized. Our study provides a first glimpse of the evolutionary developmental mechanisms driving adaptation of Phalaenopsis flowers to different pollinators by providing combined micro-morphological and molecular evidence for a possible sepaloid–petaloid–staminodial origin of the orchid mentum.

中文翻译：

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蜜蜂授粉蝴蝶兰三种高度特化花结构的进化和发展。

许多花器官的形状和大小的变化与传粉者有关。这些器官的进化是由 MADS-box 和 MYB 转录因子的复制和修饰驱动的。我们应用了微形态学（SEM 和微 3D-CT 扫描）和分子技术（转录组和 RT-PCR 分析）的组合来了解愈伤组织、stelidia 和 mentum 的进化和发展，这三种高度专业化的兰花花结构涉及在授粉。从蜜蜂授粉的蝴蝶兰和红花蝴蝶兰的花中收集早期和成熟的组织，这两种花的形态不同：P. equestris 有一个大的愈伤组织，但有短的 stelidia 并且没有mentum，而 P. pulcherrima 有一个小愈伤组织，但长的 stelidia 和明显的精神。我们的研究结果表明，stelidia 从早期的原始阶段发展而来，而愈伤组织和心理发育较晚。结合显微 3D-CT 扫描分析和基因表达分析表明，愈伤组织为混合花瓣状 - 退化角起源，退化角起源于 stelidia，以及混合萼片-花瓣-退化角起源。SEP 进化枝 1 拷贝在 P. equestris 的较大愈伤组织中表达，而 AP3 进化枝 1 和 AGL6 进化枝 1 拷贝在 P. pulcherrima 的显着的mentum 和长 stelidia 中表达。AP3 进化枝 4、PI-、AGL6 进化枝 2 和 PCF 进化枝 1 拷贝可能在愈伤组织和合股骨发育中起平衡作用。一方面，愈伤组织中 DIV 进化枝 2 表达与 SEP 进化枝 1 表达之间似乎存在权衡，并且在另一方的柱状体和脑部中具有AP3进化枝1和AGL6进化枝1表达。我们检测到了两种蝴蝶兰的愈伤组织、柄部和被膜中MADS box AP3/PI-like、AGL6-like和SEP-like以及MYB DIV-like基因拷贝的差异生长和表达，其中这些花结构非常不同的形状和大小。我们的研究通过提供组合的微观形态学和分子证据来证明兰花心部可能的萼片-花瓣-雄蕊起源，首次揭示了推动蝴蝶兰花适应不同传粉者的进化发育机制。其中这些花卉结构的形状和大小非常不同。我们的研究通过提供组合的微观形态学和分子证据来证明兰花心部可能的萼片-花瓣-雄蕊起源，首次揭示了推动蝴蝶兰花适应不同传粉者的进化发育机制。其中这些花卉结构的形状和大小非常不同。我们的研究通过提供组合的微观形态学和分子证据来证明兰花心部可能的萼片-花瓣-雄蕊起源，首次揭示了推动蝴蝶兰花适应不同传粉者的进化发育机制。