Figure 5

Junction sequences of the R2 elements with the 28S gene. (a) 3' junction sequences. All Drosophila R2 elements contain 3' poly(A) tails. Most R2 insertions are consistent with the location of the R2 DNA cleavage sites on the bottom strand (see panel (c)) and its use in priming reverse transcription. (b) Representative examples of the 5' junctions of R2 elements with the 28S gene. All full-length examples are from D. melanogaster. R2 sequences are boxed, 28S sequences are in bold, non-templated sequences are in plain text, and duplications of 28S sequences are underlined. Boxed residues shaded grey correspond to microhomologies: sequences that could correspond to either the 28S sequence or the R2 element. (c) Location of the probable cleavage sites on the 28S gene. Arrows show cleavage locations determined in vitro for the R2 endonuclease from B. mori [10]; the arrow head topped by '0' shows the location of the top-strand cleavage site inferred after analysis of the Drosophila R2 5' junctions. The bottom diagram shows a hypothetical intermediate in the integration reaction after first-strand synthesis (boxed nucleotides) and second strand cleavage. The terminal two nucleotides of the cDNA are proposed to anneal to the top strand of the cleaved target site. This microhomology allows precise priming of second-strand DNA synthesis and the generation of the precise junctions seen in example a in panel (b).