Proceedings of the XLVI Italian
Society of Agricultural Genetics - SIGA Annual Congress
Giardini
Naxos, Italy - 18/21 September, 2002
ISBN 88-900622-3-1
Poster
Abstract - 2.17
CLONING OF 2,3-OXIDOSQUALENE CYCLASE INVOLVED
IN TRITERPENOID SAPONINS PATHWAY IN ASTER SEDIFOLIUS
CAMMARERI M.*, CONSIGLIO F.**,
ZACCARDELLI M.***, ERRICO A.*, CONICELLA C.**
*) Dept. of Soil,
Plant and Environmental Sciences, University of Naples "Federico II",
Via Università, 100, 80055 Portici (NA)
**) CNR-IMOF,
Research Institute for Vegetable and Ornamental Plant Breeding, Via
Università, 133 - 80055 Portici (NA)
***) Research
Institute for Industrial Crops, MiPAF, S.S 18, 156, 84091 Battipaglia (SA)
antitumor
effect, degenerate primers, nested PCRs
Many
saponins are present in higher plants, and saponin-containing plants have been
used for different medical purpose. Recent studies on the biological activities
of pure saponins have demonstrated antiinflammatory activities, antibacterial action,
stimolation of lipid metabolism and antitumor effect. Several species of the
genus Aster (A. lingulatus, A. yunnanensis, A. batagensis, A. scaber, A. tataricus) produce triterpenoid saponins and,
particularly, asterlingulatosides A, B, C and D, isolated from A.
lingulatus, showed good
inhibitory activity against DNA synthesis in human leukemia. Many triterpenoic
aglycones of saponins are biosynthesized from a common precursor,
2,3-oxidosqualene. The cyclization of this compaund in pentacyclic carbon
skeleton is catalyzed by the 2,3-oxidosqualene cyclase.
Objective of the present work is the molecular cloning of 2,3-oxidosqualene cyclase in Aster. A homology-based PCR method was applied by designing two sets of degenerate oligonucleotide primers at the regions which are highly conserved among known oxidosqualene cyclase genes. Total RNA was extracted from leaves of Aster sedifolius at the ‘rosette’ stage and cDNA was obtained by reverse transcriptase. Nested PCRs were carried out to amplify the core fragment of the putative gene for 2,3- oxidosqualene cyclase. A fragment of 660 bp was obtained. A pPCR Script Amp vector was used for subcloning and sequencing. The sequence of core fragment showed 83% of identity with 2,3-oxidosqualene cyclase of Panax ginseng.