BUZGO, MATYAS1*, YIN-LONG QIU2, and MARK W. CHASE3. 1Department of Botany, University of Florida, Bartram, P.O. Box 118526, Gainesville, FL 32611-8526, USA; 2Biology Department, Morrill Science Center, 611 North Pleasant Street, University of Massachusetts, Amherst, MA 01003-5810, USA; 3Jodrell Laboratory, Royal Botanical Gardens, Kew, Richmond, Surrey, TW9 3DS, UK. - ITS in Arisaema (Araceae).
Arisaema (Araceae) has an unusually widespread, temperate
distribution including eastern Africa, Asia, and North America. It has
consistent basic morphological features, but a high diversity in its
specific modifications on various structures (e.g., shape and
coloration of the spathe and appendix, paradioecy, leaf divisions,
tuber shape etc.). This suggests recent speciation, and obscures
species delimitations and homologies. For this phylogenetic analysis,
42 taxa were sampled (incl. Pinellia, Arisarum and
Biarum), represented by a total of 74 DNA sequences of ITS1,
ITS2, and the 5.8S region of the nuclear rRNA genes.
Amplification with primers specific for the ITS region of
plants turned out to be difficult. By modifying the protocols, cloning
and trying different accessions of the same species, potential
problems were evaluated, such as degeneration of DNA, inhibition by
secondary compounds or polysaccharides, and effects caused by
secondary structure. Only the use of less specific primers resulted in
good PCR yields. This implies that the polymerase is not inhibited,
but rather that the primer binding site may be inaccessible for
specific primers due to strong, species-specific secondary structures
(or proteins). The sequences obtained represent paralogs that are
difficult to align; hence, relationships could not be resolved. The
activity of the paralogs could not be determined. Possible ways to
overcome this primer-binding problem, to select active or homologous
ITS sequences, and the potential of ITS-paralogs for the
study of recent speciation are discussed.
Key words: Arisaema Araceae ITS Secondary Structure