A genomic clone encoding a phospholipid transfer protein from barley.

A barley (Hordeum vulgare cv Igri) genomic library was screened with synthetic oligonucleotides that correspond to the noncoding sequences of a barley PLTP cDNA (Molina and García-Olmedo, 1993). Plaque hybridizations were performed at moderate stringency with 32P-end-labeled oligonucleotides. Nitrocellulose filters were exposed to film for 16 h at -80°C. A positive plaque was isolated, and the genomic insert was analyzed by Southern hybridization. A 2.1-kb NotIHindIII fragment that hybridized to a maize PLTP cDNA probe was subcloned into pGEM 11Zf(+). Sequencing of this fragment indicated that it contained an open reading frame of 345 bp that encodes a PLTP. The predicted molecular mass of the mature protein is 8.7 kD with a pI of 8.85. It has a 25-amho acid residue signal sequence based on alignment with other PLTP sequences. The deduced amino acid sequence of the PLTP precursor encoded by this genomic clone is very closely related to that of pKG285 (95% identical), a barley PLTP cDNA previously reported to be expressed in barley leaves and coleoptiles (Gausing, 1994). It is also related to LTP2 (75% identical), LTP3 (72% identical), and LTP4 (88% identical) from barley leaves (Molina and García-Olmedo, 1993) but less related to both the cold-induced BLT4 (43% identical) from barley meristematic tissue (Dunn et al., 1991) and the probable amylase/ protease inhibitor (43% identical) from barley aleurone (Mundy and Rogers, 1986). Based on these homologies, we believe that the regulatory sequences contained in this genomic clone will be able to direct gene expression in the epidermal cells of barley leaves, perhaps in a pathogen-inducible manner (Molina and García-Olmedo, 1993). The 5' nontranslated sequence contains a potential TATA box 78 bp upstream of the initiator methionine codon and the 3' nontranslated sequence contains a polyadenylation signal 115 bp beyond the stop codon. There is no interruption of the protein-coding sequence by introns.