| Summary: | Slipper orchids (subfamily Cypripedioideae) comprise five genera; Paphiopedilum, Cypripedium, Phragmipedium, Selenipedium, and Mexipedium. Phylogenetic relationships of the genus Paphiopedilum, were studied using nuclear ribosomal ITS and plastid sequence data. The results confirm that Paphiopedilum is monophyletic and support the division of the genus into three subgenera Parvisepalum, Brachypetalum and Paphiopedilum. Four sections of subgenus Paphiop edilum (Pardalopetalum, Cochlopetalum, Paphiopedilum and Barbata) are recovered with strong support for monophyly, concurring with a recent infrageneric treatment. Section Coryopedilum is also recovered with low bootstrap but high posterior probability values. Relationships in Barbata remain unresolved, with short branch lengths and narrow geographical distributions suggesting it may have undergone rapid radiation. Genome sizes were measured for seven taxa in Paphiopedilum and chromosome and genome size data mapped onto the phylogenetic framework, showing no clear trend in increase in chromosome number in the genus. The diploid chromosome number of 2n = 26 in subgenera Parvisepalum and Brachypetalum suggests it is the ancestral condition, with higher chromosome numbers in Cochlopetalum and Barbata pointing to centric fission possibly having occurred independently in these sections. Although species in Barbata have larger genome sizes than other sections, any trend of genome size evolution remains unclear in the genus. Eight primer pairs for plastid microsatellites were designed from consensus sequences generated from different genera, most of them shown to be applicable across the subfamily. High levels of variation in allele size were observed at interspecific levels but at intraspecific level, low levels were observed in Cypripedium calceolus. The application of plastid microsatellites for population genetic analyses in C. calceolus was limited because few of them are polymorphic and low numbers of alleles were detected. Results were generally congruent with a previous study. Within the limits of this data, the plastid haplotype distribution of C. calceolus in western and northern Europe could indicate possible recolonisation routes from three main refugia, following glaciations. Size variation has also been detected in other species in some markers but sampling was sparse.
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