UNIVERSAL PCR PRIMERS FOR DETECTION OF PHYTOPATHOGENIC AGROBACTERIUM STRAINS

Two PCR primer pairs, based on the virD2 and ipt genes, detected a wid e variety of pathogenic Agrobacterium strains. The endonuclease domain of VirD2 protein, which cleaves transferred DNA (T-DNA) border sequen ces, is highly conserved; primer oligonucleotides specific for the end onuclease portion of virD2 detected all pathogenic strains of Agrobact erium tested. PCR primers corresponding to conserved sequences in ipt, the T-DNA-borne cytokinin synthesis gene, detected only Agrobacterium tumefaciens and distinguished it from Agrobacterium rhizogenes. The v irD2 and ipt primer pairs did not interfere with each other when inclu ded in the same PCR amplification, and this permitted simultaneous det ection of both genes in a single reaction. One nonpathogenic Agrobacte rium radiobacter strain contained virD2 but not ipt; we speculate that this strain arose from a pathogenic progenitor through a deletion in the T-DNA. The virD2 primer pair appears to be universal for all patho genic Agrobacterium species; used together, the primer sets reported h ere should allow unambiguous identification of Ti plasmid DNA in bacte ria isolated from soil and plants.