The tomato polygalacturonase gene and ripening-specific expression in transgenic plants

Abstract

Polygalacturonase (PG) is the major cell wall degrading enzyme of tomato fruit. It is developmentally regulated and is synthesised de novo in ripening fruit. Genomic clones encoding a PG gene of tomato (Lycopersicon esculentum Mill cv. Ailsa Craig) have been isolated, mapped and sequenced. The sequence of the protein-coding region is identical to that of a PG cDNA [20]. Comparison of the cloned restriction fragments with genomic Southern data suggests that there may only be one gene for PG per haploid genome. The PG gene, which covers approximately 7 kb, is interrupted by 8 intervening sequences ranging in size from 99 bp to 953 bp. The transcription start point was identified by S1 mapping and primer extension analysis. About 1.4 kb of 5' flanking DNA has been sequenced. This contains putative TATA and CAAT boxes and also direct repeat sequences. A transcriptional fusion has been constructed between the putative 1.4 kb promoter fragment and the chloramphenicol acetyl transferase (CAT) gene. Constructs containing this gene have been transferred to tomato using binary vectors. Regenerated transgenic plants express CAT in ripe tomato fruit, but not in unripe tomatoes, leaves, or roots.