Cloning Of The Allene Oxide Systhase Gene From Tomato

Abstract

A tomato mutant called JL5 is defective in the wound signaling cascade leading to defense against insects. A deficiency in allene oxide synthase (AOS), an enzyme involved in the biosynthesis of jasmonic acid, has been suggested as the cause of this defect. In an effort to test this hypothesis, we attempted to clone the AOS gene from tomato. Two strategies were used to achieve this goal. In the first cloning strategy, Polymerase Chain Reaction (PCR) experiments were performed using oligonucleotide primers that were designed from a comparison of the published flax and rubber tree AOS cDNA sequences. These experiments revealed amplified DNA fragments from a tomato cDNA library template of the predicted size. In the second cloning strategy, an AOS cDNA clone from Arabidopsis thaliana (At) was used to screen a tomato cDNA library. Ten positive plaques from a primary screen were purified. DNA sequence analysis revealed a full length clone (called AOS1) encoding a polypeptide of 510 amino acids. Comparison of this sequence with those in protein sequence databases suggests that AOS1 encodes an allene oxide synthase.