Identification and phylogenetic analysis of gypsy-type retrotransposons in the plant kingdom.

PCR was performed with degenerate primers which hybridized to the homologous sequences in the reverse transcriptase (rt) genes of gypsy-type retrotransposons from rice (RIRE3, RIRE8 and RIRE2), using total DNA samples from various plants (monocots, dicots, pine, ginkgo, horsetail, liverwort and algae) as templates. Cloning and sequencing showed that the amplified fragments had various degrees of homology to the rt sequences of rice retrotransposons. Phylogenetic analysis showed that these retrotransposon homologues and some additional gypsy-type retrotransposons previously identified from plants could be classified into two families, A and B. In each family, the retrotransposons were further classifiable into several subfamilies. Interestingly, retrotransposons from a single or related plant species were clustered in each subfamily. This indicates that sequence divergence during vertical transmission has been a major influence on the evolution of gypsy-type retrotransposons in plants. The retrotransposons isolated from one plant species could often be classified into the two families. This indicates that the gypsy-type retrotransposons of a family evolved independently within a species without affecting the evolution of retrotransposons of the other family. Retrotransposons in each subfamily are characterized by the lengths of LTR, by the nucleotide sequences in the terminal regions of LTRs, and by the PBS (primer binding site) sequence complementary to the 3' sequence of a particular tRNA species.