Rhynchostylis retusa (L.) Blume is a popular and scarce orchid species with attractive flowers arranged in a racemose cluster. To study this flower’s complex floral perianth formation, MADS-box genes that play an important role in flower morphogenesis were investigated. Four B-class DEF-like MADS-box genes, RrDEF1, RrDEF2, RrDEF3, and RrDEF4, and two E-class AGL6-like MADS-box genes, RrAGL6-1 and RrAGL6-2, were identified, and their expression was characterized by qRT-PCR. These genes were expressed in every developmental stage of reproductive organs but not in the vegetative leaves, except for RrDEF2. Most of these genes revealed similar expression patterns in stage 3 and mature flowers. Higher RrDEF1 and RrAGL6-1 expression was detected in the sepals and petals rather than the lips, while RrDEF3, RrDEF4, and RrAGL6-2 were highly expressed in the lips but exhibited low expression in the sepals. RrDEF3 was also strongly expressed in petals. These gene expression patterns supported current profiles involving sepal/petal/lip determination, with RrDEF1 and RrAGL6-1 promoting sepal/petal formation, while RrDEF3, RrDEF4, and RrAGL6-2 promoted lip formation. Our findings support revised ‘orchid code’ and ‘perianth code’ hypotheses in which floral organs are regulated by combined levels of expression from each clade of DEF-like genes and AGL6-like genes. These four RrDEF and two RrAGL6 genes promote and enrich our knowledge and understanding concerning the floral development of orchids.

Rhynchostylis retusa（L.）Blume是一种流行而稀少的兰花物种，其美丽的花朵排列在消旋簇中。为了研究该花的复杂花被膜形成，研究了在花朵形态发生中起重要作用的MADS-box基因。鉴定了四个B类DEF样MADS-box基因RrDEF1，RrDEF2，RrDEF3和RrDEF4，以及两个E类AGL6样MADS-box基因RrAGL6-1和RrAGL6-2，并对其表达进行了表征通过qRT-PCR。这些基因在生殖器官的每个发育阶段均表达，但在营养叶中不表达，RrDEF2除外。这些基因中的大多数在第3阶段和成熟花中显示出相似的表达模式。在萼片和花瓣而不是嘴唇中检测到更高的RrDEF1和RrAGL6-1表达，而RrDEF3，RrDEF4和RrAGL6-2在嘴唇中高表达，但在萼片中低表达。RrDEF3在花瓣中也强烈表达。这些基因表达模式支持涉及萼片/花瓣/嘴唇测定的当前概况，其中RrDEF1和RrAGL6-1促进萼片/花瓣形成，而RrDEF3，RrDEF4和RrAGL6-2促进唇部形成。我们的发现支持修正的“兰花代码”和“花粉代码”假说，在这些假说中，花器官受DEF样基因和AGL6样基因各分支的表达水平的共同调节。这四个RrDEF和两个RrAGL6基因促进和丰富了我们对兰花花发育的知识和了解。