Detection and Investigation of Genetic Relatedness among Aster Yellows and Other Mycoplasmalike Organisms by Using Cloned DNA and RNA Probes. I. -M. Lee. Microbiology and Plant Pathology Laboratory, Agricultural Research Service, United States Department of Agriculture, Beltsville, Maryland 20705 U.S.A.. R. E. Davis. Microbiology and Plant Pathology Laboratory, Agricultural Research Service, United States Department of Agriculture, Beltsville, Maryland 20705 U.S.A.. MPMI 1:303-310. Accepted 27 October 1988. This article is in the public domain and not copyrightable. It may be freely reprinted with customary crediting of the source. The American Phytopathological Society, 1988.

A method was developed for enriching the concentration of mycoplasmalike organism (MLO) DNA in diseased plant extracts. With this method, fragments of DNA of the aster yellows (AY) MLO were cloned in pSP6 plasmid vectors and amplified in Escherichia coli strain JM83. Labeled double-stranded DNA probes were prepared by nick translation of recombinant plasmids by using either ³²P nucleotides or biotinylated nucleotides. Single-stranded RNA probes (riboprobes) were synthesized in vitro by using SP6 polymerase and linearized recombinant plasmids and were labeled with ³²P. The probes all hybridized with nucleic acid from AY MLO-infected, but not healthy, plants. Some hybridized also with nucleic acid from plants infected by other MLOs. Hybridization patterns indicated existence of a cluster of MLOs, including AY MLO, that share greater nucleotide sequence homology with one another than with other MLOs. Riboprobes were used to differentiate AY MLO from others in the AY MLO cluster.