A coupled expression system for plants was established in this study. The 5’-terminal of T7 RNA poly-merase gene was modified by addition of the coding sequence of nuclear location signal from SV40 large T antigen. Plant expression vector pBBT7 was constructed with the modified T7 RNA polymerase gene under the control of CaMV35S promoter. Another expression vector pBTG contained cassette of gusA controlled by T7 promoter. The two vectors were co-transformed into tobacco via the Agrobecte-rium -mediated method. Results of GUS activity indicated that the co-transformed plant with pBBT7 and pBTG showed a high level of GUS activity. The results demonstrated that the coupled expression system of T7 polymerase and T7 promoter was workable in plants.
A novel cDNA sequencehtMT2, which encodes a type 2 metallothionein_like protein, was isolated from Helianthus tuberosus L. tuber cDNA library. The whole sequence is 509 bp, including an open reading frame (ORF) of 240 bp, a 5′ UTR of 62 bp and a 3′ UTR of 207 bp. Two genomic sequences covering the coding region ofhtMT2were cloned by PCR reaction. Sequence analysis revealed that the genomic sequences htMTG_1 of 986 bp and htMTG_2 of 982 bp were both composed of three exons and two introns. The deduced protein consisted of 79 amino acid residues with a predicted molecular weight of 7.8 ku (kD). Amino_terminal and carboxy_terminal domains contained 8 and 7 cysteine residues respectively, separated by a central cysteine free spacer. Sequence alignment revealed that the predicted protein ofhtMT2 was homologous to type 2 metallothioneins (MTs) of plants. Southern blotting analysis indicated that htMT2was encoded by a small multi_gene family in H. tuberosus genome. Northern blotting analysis showed that htMT2 transcripts were detected in stems, leaves and leafstalks, but no transcripts were detected in roots. The expression level in stems was the highest among the above tissues. Transcripts in stems were significantly reduced by Cu 2+ treatment. Judging from the homologies between the deduced HtMT2 and other type 2 plant metallothioneins as well as responses to metal ions, we believe thatwere cloned by PCR reaction. Sequence analysis revealed that the genomic sequences htMTG_1 of 986 bp and htMTG_2 of 982 bp were both composed of three exons and two introns. The deduced protein consisted of 79 amino acid residues with a predicted molecular weight of 7.8 ku (kD). Amino_terminal and carboxy_terminal domains contained 8 and 7 cysteine residues respectively, separated by a central cysteine free spacer. Sequence alignment revealed that the predicted protein ofhtMT2 was homologous to type 2 metallothioneins (MTs) of plants. Southern blotting analysis indicated that htMT2was encoded by a small multi_gene family in H. tube
