CONSTRUCTION OF NICOTIANA-TABACUM-L TRANS GENIC PLANTS EXPRESSING THEBACTERIAL GENE FOR BETA-1,3-GLUCANASE .1. CONSTRUCTION OF VECTORS FORTRANSFECTION OF PLANT-CELLS AND EXPRESSION OF THE MODIFIED BETA-1,3-GLUCANASE GENE FROM CLOSTRIDIUM-THERMOCELLUM IN TOBACCO PROTOPLASTS

We constructed two vectors, pC27-glc and pC29-glc, that allow expressi on of the beta-1,3-glucanase gene (glc) in plant cells. The glc gene w as previously cloned from anaerobic thermophilous bacterium Clostridiu m thermocellum. To increase the efficiency of expression, the N-termin al fragment of the glc gene encoding bacterial transient peptide was d eleted, and hybrid variants of lacZ-glc were obtained. Analysis of exp ression of the hybrid genes in Escherichia coli showed that deletion o f the fragment corresponding to 31 amino acids (a.a.) of beta-glucanas e affected neither activity nor thermostability of the enzyme. The mod ified gene was subcloned into two vectors, pC27 and pC29, in which its expression was controlled by the T(R)2' promoter pf the 2' gene of T- DNA and the rbcS promoter from Arabidopsis, respectively. Each of the resulting plasmids, pC27-glc and pC29-glc, was transfected into protop lasts of Nicotiana plumbaginifolia. Both the plasmids were shown to al low a high level of activity of the thermostable beta-1,3-glucanase. W e plan to use the vectors obtained for transformation of agrobacteria and construction of transgenic plants.