ISOLATION OF43 MICROSATELLITE LOCI FROM PAPHIOPEDILUMROTHSCHILDIANUM, AN ENDANGERED SPECIES OF SLIPPER ORCHID

摘要

Paphiopedilum mthschildiamtm is one of the endangered species of slipper orchids associated with specific rnicrohabitats within Sabah, The genomic characterization and subsequent assessment of interspecific genetic variability of P, mthschildmnum willprovide a valid basis for the determination of those genetic factors that are significant for the survival of a species classified as endangered. MicrosatelHtes are significant molecular markers in the context of population genetic studies as they can be employed to determine interspecific and Intraspecific genetic variability. This variability can then be utilized to characterize the genetic resilience of a particular species when subjected to environmental stresses. Leaf tissue was sampled randomly from mature P. rathschildianum plants at the ex-situ conservation site managed by the Sabah Parks Authority at Janabalu National Park, Sabah, DNA was extracted, purified and amplified using the 5 anchored Polymerase Chain Reaction with a degenerate primerincorporating a dinucleotide repeat motif. The PCR products were cloned into a pCR 2,1 TOPO TA cloning vector and subsequently chemically transformed in E. coli competent cells, A total of 155 positive clones were obtained of which 19 were sequenced. The sequence data yielded 43 unique microsatellite loci. Investigations are now being directed toward obtaining a multiplicity of unique sequences meorporatittg microsatellite loci with di-, tri- and tetranucleotide repeat motifs. This sequence data will subsequently be utilized to design specific primers that will be employed for genetic profiling of the slipper orchid.