Genetic studies of Cypripedium calceolus (the lady’s slipper orchid)

Investigating population genetics in the lady’s slipper orchid to inform the reintroduction programme for this threatened species.

Lady's slipper orchid

The species

Unlike many Cypripedium species, our native lady’s slipper orchid (C. calceolus) is a widespread species in temperate Eurasia. However, it has been brought to the brink of extinction in many localities, including those in northern England, where it was reduced to a single site.

The recovery programme

By 1970, it was clear that positive action was required if C. calceolus was to survive in England and extensive work was initiated in situ and ex situ, under the supervision of the Cypripedium Committee, a body of interested parties led by Natural England. Work at the wild site slowly paid dividends, but the restriction of the species to just one site would always render it vulnerable to stochastic events, and reintroduction has been the subject of a long-running and high-profile conservation programme. Plants of known English origin also exist in cultivation, and self-pollination and cross-pollination of the wild individual and these plants with each other produces seeds that are germinated at Kew and result in a stock of plants with increased genetic diversity. The cross-pollinations are informed by genetic data about the relationships between the plants.

Kew’s role in the project

The seedlings for the project have all been produced using in vitro techniques at Kew, and the introduction of artificially propagated seedlings across the former range in northern England had led to flowering at 11 sites in England by 2014. In addition, we were asked to provide genetic data in support of this reintroduction programme and information on the relationship of English plants to populations elsewhere in Europe. This initially proved more difficult than expected due to the large genome size in this species; we showed that this made some of the standard genetic fingerprinting techniques available at the time difficult or impossible to apply to this species and others with large genomes. As a result of the development of new techniques, based on next-generation sequencing, we have now overcome these problems for C. calceolus and we have produced detailed fingerprints from the English plants and representative plants from elsewhere in Europe. These are being used to direct the reintroduction programme.


  • To develop a system for genetic fingerprinting of Cypripedium calceolus, overcoming problems associated with large genome size.
  • To fingerprint all native plants of Cypripedium calceolus and representative plants from elsewhere in Europe.
  • To provide genetic data for Cypripedium calceolus in support of the reintroduction programme in England.
Project Leader

Michael F. Fay


Roberta Gargiulo

  • A set of nuclear and plastid microsatellites for Cypripedium calceolus.
  • Genotypes of native plants and of seedlings of Cypripedium calceolus to be used in reintroduction as required by Natural England.
  • Clarification of the relationship of the English populations of Cypripedium calceolus to populations elsewhere in Europe.


  • Natural England


  • University of Copenhagen, Denmark
  • University of Tartu, Estonia
  • University of Helsinki, Finland

Natural England

Fay, M. F. & Taylor, I. (2015)

801. Cypripedium calceolus. Orchidaceae

Curtis’s Botanical Magazine 32: 24-32

Pedersen, H. Æ., Rasmussen, H. N., Kahandawala, I. M. & Fay, M. F. (2012)

Genetic diversity, compatibility patterns and seed quality in isolated populations of Cypripedium calceolus(Orchidaceae)

Conservation Genetics 13: 89-98

Fay, M. F., Bone, R., Cook, P., Kahandawala, I., Greensmith, J., Harris, S., Pedersen, H. Æ., Ingrouille, M. J. & Lexer, C. (2009)

Genetic diversity in Cypripedium calceolus (Orchidaceae) with a focus on northwestern Europe, as revealed by plastid DNA length polymorphisms

Annals of Botany 104: 517-525

Fay, M. F., Cowan, R. S. & Leitch, I. J. (2005)

The effects of nuclear DNA content (C-value) on the quality and utility of AFLP fingerprints

Annals of Botany 95: 237-246

Fay, M. F. & Cowan, R. S. (2001)

Plastid microsatellites in Cypripedium calceolus(Orchidaceae): genetic fingerprints from herbarium specimens

Lindleyana 16: 151-156