Non-LTR retrotransposons (LINEs) as ubiquitous components of plant genomes

During the course of work aimed at isolating a rice gene from Oryza austral iensis by PCR, the oligonucleotide primers used were found to generate a fr agment that showed sequence homology to the endonuclease (EN) region of the maize non-LTR retrotransposon (LINE) Cin4. We carried out further PCRs usi ng oligonucleotide primers that hybridized to these sequences, and found th at they amplified several fragments, each with homology to the EN regions, from Oryza sativa cv. Nipponbare as well as O. australiensis. We mapped the approximate locations of two rice LINE homologues by screening clones in a YAC library made from a rice (O. sativa) genome, and found that each homol ogue was present in a low copy number apparently at nonspecific regions on rice chromosomes. We then carried out PCR using degenerate oligonucleotide primers which hybridized to the rice LINE homologues and Cin4 to ascertain whether LINE homologues are present in a variety of members of the plant ki ngdom, including angiosperms, gymnosperms, bracken, horsetail and liverwort . Cloning and nucleotide sequencing revealed that 53 clones obtained from 2 7 out of 33 plant species contained LINE homologues. In addition to these h omologues, we identified four homologues with EN regions in the Arabidopsis thaliana genome by a computer search of databases. The nucleotide sequence s of almost all the LINE homologues were greatly diverged, but the derived amino acid sequences were well conserved, and all contained glutamic acid a nd tyrosine residues at almost the same relative positions as in the the ac tive site regions of AP (apurinic/apyrimidinic) endonucleases. The EN regio ns in the LINE homologues from closely related plant species show a closer phylogenetic relationship, indicating that sequence divergence during Verti cal transmission has been a major influence upon the evolution of plant LIN Es.