Fig. 1: Insertion of the LINE-1 (L1) retrotransposon and its detection by TotalReCall.

a Haploid copy of the genome before retrotransposition. b Endonuclease break in each strand of DNA. c L1 RNA directly reverse transcribed into the genome resulting in the synthesis of the single-stranded cDNA starting from the 3’ end of the L1 transcript and extending a variable length towards its 5’ end. Teal, L1 RNA. Green, reverse transcribed poly(T) cDNA. Blue, reverse transcribed L1 cDNA. d Twin priming results in the simultaneous reverse transcription of different parts of the L1 transcript into the two strands of the genome. Red, reverse transcribed L1 cDNA on the opposite gDNA strand. e, f Genome after synthesis of the second strand of DNA and repair. Components of the L1 sequence are annotated with respect to the “top” strand of the genome. Purple, target site, which is duplicated following repair. Red, newly inserted L1 sequence that was synthesized on the top strand and is therefore reverse complemented with respect to L1 RNA. Blue, newly inserted L1 sequence that was synthesized on the bottom strand. Green, newly inserted poly(A). Paired-end reads originating from the modified genomes are shown. e The process shown in c results in a (possibly 5’ truncated) “canonical” retrotransposition. f The process shown in d results in an “inversion-containing” retrotransposition. The resulting genomic sequence has two L1 fragments in opposite orientations. g Mapping of reads A–D to the unmodified (reference) genome lacking the transposon insertion (left) and the transposon sequence (right). Left, tails of reads A1, B1, and D1 that come from the novel transposon are clipped (shown as dashed lines). Right, read C2 and the clipped tails of reads A1 and D1 align to the transposon sequence. The clipped tail of read B1 contains only poly(T). In the absence of inversion, the alignment between the clipped sequence and the transposon sequence reflects the length of the newly inserted transposon. When inversion occurs, such an alignment will only reflect the position where the second priming occurred.