Genetic differentiation of phytoplasma isolates by DNA heteroduplex mobility assay and single-strand conformation polymorphism analysis

Heteroduplex mobility assay (HMA) and single-strand conformation polymorphism (SSCP) analyses combined with PCR were developed for genetic differentiation of various phytoplasma isolates. In the HMA and SSCP analyses, differences in the mobility shifts and the SSCP band patterns identified three distinct types of phytoplasmas: Type I, jujube witches'-broom (JWB) and ligustrum witches'-broom (LiWB); Type II, mulberry dwarf (MD) and sumac witches'-broom (SuWB); and Type ID, paulownia witches'-broom (PaWB). Results of the sequence analyses revealed that phytoplasmas of JWB and MD had 100% homology with LiWB and SuWB, respectively. On the other hand, PaWB phytoplasma had 97.8% homology with MD phytoplasma. The PCR-HMA and SSCP techniques were very useful in determining variations in sequence among several isolates of phytoplasmas. Furthermore, the methods were rapid, economical, highly sensitive, and easy to handle with the gels.