S Tegli1, A Sereni, G Surico. 1. Dipartimento di Biotecnologie Agrarie (DiBA) - Sezione di Patologia Vegetale, Università degli Studi di Firenze, Firenze, Italy. stefania.tegli@unifi.it
Abstract
AIMS: To develop a PCR-based protocol for the rapid, sensitive and specific detection of Curtobacterium flaccumfaciens pv. flaccumfaciens (Cff) in bean seeds. METHODS AND RESULTS: A pair of PCR primers (CffFOR2-CffREV4), targeting the sequence of a cloned DNA fragment of 550 bp amplified in Repetitive-sequence-based-PCR (Rep-PCR) experiments, were designed and shown to specifically amplify a 306-bp DNA fragment using Cff DNA as template. Moreover, this PCR protocol was demonstrated to successfully detect Cff in naturally infected bean seeds in 36 h. CONCLUSIONS: A specific, highly sensitive and rapid PCR assay for the detection of Cff was achieved. SIGNIFICANCE AND IMPACT OF THE STUDY: Cff is a seed-borne bacterium on the EPPO A2 quarantine list; this procedure may be useful for routine diagnosis of Cff, overcoming the problems of conventional techniques.
AIMS: To develop a PCR-based protocol for the rapid, sensitive and specific detection of Curtobacterium flaccumfaciens pv. flaccumfaciens (Cff) in bean seeds. METHODS AND RESULTS: A pair of PCR primers (CffFOR2-CffREV4), targeting the sequence of a cloned DNA fragment of 550 bp amplified in Repetitive-sequence-based-PCR (Rep-PCR) experiments, were designed and shown to specifically amplify a 306-bp DNA fragment using Cff DNA as template. Moreover, this PCR protocol was demonstrated to successfully detect Cff in naturally infected bean seeds in 36 h. CONCLUSIONS: A specific, highly sensitive and rapid PCR assay for the detection of Cff was achieved. SIGNIFICANCE AND IMPACT OF THE STUDY: Cff is a seed-borne bacterium on the EPPO A2 quarantine list; this procedure may be useful for routine diagnosis of Cff, overcoming the problems of conventional techniques.