Transformation of white poplar (Populus alba L.) with a novel Arabidopsis thaliana cysteine proteinase inhibitor and analysis of insect pest resistance

Abstract

Transgenic white poplar (Populus alba L.) plants expressing a novel Arabidopsis thaliana cysteine proteinase inhibitor (Atcys) gene have been produced using Agrobacterium tumefaciens -mediated gene transfer. Internodal stem segments of cv. Villafranca were co-cultivated with the EHA105 pBI-Atcys A. tumefaciens strain. Sixteen putative transgenic plant lines were regenerated from different calli with a transformation efficiency of 11%. The integration and expression of the cysteine proteinase inhibitor (Atcys) gene into the plant genome was confirmed by Southern and northern blot analyses. Papain inhibitory activity was detected in poplar transgenic tissues by means of a specific in vitro assay. Such activity was sufficient to inhibit most of the digestive proteinase activity of chrysomelid beetle (Chrysomela populiL.) and confer resistance to C. populi larvae on selected transgenic plants. A close correspondence between the inhibition of papain and resistance to poplar leaf beetle was observed in all tested transgenic lines. Our results indicate that Atcys could be succesfully employed in breeding programmes aimed at the selection of new poplar genotypes resistant to major insect pests. Abbreviations: BAP ‐ 6-benzyl-aminopurine, CIM ‐ callus inducing medium, IBA ‐ indole-3-butyric acid, LB ‐ Luria-Bertani, MS ‐ Murashige & Skoog 1962, TDZ ‐ Thidiazuron, WPM ‐ woody plant medium