Molecular cloning of two different ACC synthase PCR fragments in carnation flowers and organ-specific expression of the corresponding genes

Abstract

Degenerate oligonucleotides to highly conserved regions of 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (EC 4.4.1.14), the key enzyme in ethylene biosynthesis, were used to prime the synthesis and amplification of fragments of about 1,180 bp by polymerase chain reaction (PCR) in samples of cDNA to total RNA isolated from senescing carnation (Dianthus caryophyllus) flowers. Two putative ACC synthase PCR clones were isolated one of which was identical to the sequence of a carnation ACC synthase cDNA clone (CARACC3) recently isolated by Park et al. (Plant Mol Biol 18 (1992) 377-386). The other clone (CARAS1) was ca. 66% homologous at the amino acid level to CARACC3. For both ACC synthase clones, specific oligonucleotides were synthesized and, using PCR, we were able to distinguish between the two ACC synthase transcripts in samples of total RNA isolated from different carnation flower parts and leaves. DNA blots of PCR fragments revealed that, in flowers, both ageing and ethylene stimulated the occurrence of these transcripts in an organ-specific way. CARACC3 was more abundant in RNA from the petals whereas CARAS1 was more abundant in RNA from the styles. Despite a high ethylene production observed in ovaries, the level of both transcripts was low, suggesting the existence of a third ACC synthase gene that is specifically expressed in the ovary. Transcript levels in leaves were low irrespective of treatment.