Proceedings of the XLV Italian Society of Agricultural Genetics - SIGA Annual Congress

Salsomaggiore Terme, Italy - 26/29 September, 2001

ISBN 88-900622-1-5

Poster Abstract

MOLECULAR CHARACTERIZATION OF THE TRYPSIN INHIBITOR MSTI FROM SEEDS OF SNAIL MEDIC (MEDICAGO SCUTELLATA)

CONFALONIERI M.*, BALESTRAZZI A.**, GREGUOLDO S.**, TAVA A.*, ODOARD M.*, CARBONERA D.**

* Istituto Sperimentale Colture Foraggere, Viale Piacenza 29, I-26900 Lodi

iscf@telware.it

** Dipartimento di Genetica e Microbiologia “A. Buzzati-Traverso”, Via Abbiategrasso 207, I-27100 Pavia

carbo@ipvgen.unipv.it

Medicago scutellata, proteinase inhibitors, insect resistance

Medicago scutellata or snail medic represents a commercially relevant species in Australia where it is widely used as a pasture legume. In a previous work (Ceciliani et al., 1997) the primary structure of a proteinase inhibitor named MsTI, isolated from crude seed extracts of snail medic, has been reported. MsTI is a serine-proteinase inhibitor of the Bowman-Birk family showing a high degree of identity (81%) compared to the wound-induced alfalfa trypsin inhibitor (ATI) isolated from Medicago sativa leaves (Brown et al., 1985). Moreover the insecticidal activity of MsTI has been demonstrated using digestive enzymes prepared from larvae of the insect pests Hyphantria cunea and Ostrinia nubilalis. From this point of view, Medicago scutellata might represent a useful source of insect-resistant germplasm. We are currently involved in the molecular characterization of MsTI. Preliminary northern experiments, performed using the alfalfa trypsin inhibitor ATI as a molecular probe and RNA poly(A)⁺ prepared from immature seeds of Medicago scutellata have demonstrated the presence of a 0.5 kb transcript, while Southern analysis suggested the presence of a multigene family. In order to clone the MsTI gene we have undertaken the construction of a cDNA library using RNA poly(A)⁺ extracted from immature seeds of snail medic; subsequently this library is being screened with the ATI cDNA as molecular probe. We are also evaluating the distribution of the MsTI transcript in different plant tissues by northern analysis using the alfalfa heterologous probe. The accumulation of MsTI mRNA in response to wounding and following exposure to salycilic acid and jasmonate is also being monitored. As a parallel approach, polyclonal antibodies directed against the MsTI protein have been produced and are now used in western blot experiments to characterize the expression pattern at the protein level.

References

Ceciliani F., Tava A., Iori R., Mortarino M., Odoardi M., Ronchi S. Phytochemistry 44, 393-398 (1997)

Brown W.E., Takio K., Titani K., Ryan A.C. Biochemistry 24, 2105 (1985)