Molecular cloning of a genomic DNA encoding the potato β-1,3-glucanase

A genomic clone for potato β-1,3-glucanase was isolated to study the regulatory expression mechanism of the plant gene. A genomic library was constructed with total DNA isolated from potato (Solanum tuberosum L. cv. Sumi) leaves. The genomic library was screened by plaque hybridization using the cDNA clone for tobacco β-1,3-glucanase as a probe. The genomic clone, λGlc1, for β-1,3-glucanase was isolated from the library mapped by restriction analysis using Southern hybridization. The Southern blot analysis showed that a 2.3 kb fragment contained the coding region for β-1,3-glucanase. The structure of the genomic clone was determined by nucleotide sequencing, which, together with RNA mapping with nuclease S1, showed that the cloned β-1,3-glucanase gene (λGluB3) was a genomic counterpart of the β-1,3-glucanase gene (GluB3) from a different potato cultivar (S. tuberosum L. cv. Datura) and consisted of two exons and one intron encoding a protein of 315 amino acid residues. The major transcription initiation site of λGluB3 was determined by primer extension and it appeared to be at 27 bp upstream of the translation initiation site. The canonical TATA box and AGC-enhancer elements were found in the promoter region of λGluB3 and sequence comparison revealed that the relevant promoter region of λGluB3 was more similar to that of tobacco than that office or barley.