Sequence and expression of a stem-abundant caffeic acid O-methyltransferase cDNA from perennial ryegrass (Lolium perenne)

Abstract

A perennial ryegrass (Lolium perenne) cDNA library was screened with a PCR-amplified comt DNA fragment generated from ryegrass cDNA template using degenerate oligonucleotide primers. A full-length cDNA (LpeComt1) was isolated and shown to encode a caffeic acid O-methyltransferase (COMT) enzyme by analysis of its predicted amino acid sequence. This cDNA was expressed in E.coli and COMT activity was demonstrated in bacterial extracts. The predicted amino acid sequence of the perennial ryegrass COMT was highly similar (88%) with that of a COMT isolated from maize. Southern blot hybridisation analysis using this cDNA indicated that it is a member of a small gene family comprising at least two genes in perennial ryegrass. The LpeComt1 gene is strongly expressed in stem tissue, but not obviously in shoot, leaf blade, leaf sheath, flower or root tissue.