Serial number tagging reveals a prominent sequence preference of retrotransposon integration.
Bottom Line: To address this problem we developed the serial number system, a TE tagging method that measures the frequency of integration at single nucleotide positions.We sequenced 1 million insertions of retrotransposon Tf1 in the genome of Schizosaccharomyces pombe and obtained the first profile of integration with frequencies for each individual position.Integration levels at individual nucleotides varied over two orders of magnitude and revealed that sequence recognition plays a key role in positioning integration.
Affiliation: Section on Eukaryotic Transposable Elements, Program in Cellular Regulation and Metabolism, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA.Show MeSH
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Mentions: To study genome-wide integration, a copy of Tf1 marked with neo (Tf1-neo) is expressed in S. pombe and cells with insertions are selected on media containing G418 (18,19). High-throughput sequencing of Tf1-neo integration sites relies on amplifying insertions with ligation-mediated PCR (4). The previous study that demonstrated Tf1-neo integrates into pol II transcribed promoters measured the number of positions with integration. However, the number of insertions at each position could not be determined; the duplicate sequence reads were discarded because those generated by independent integration could not be distinguished from ones resulting from PCR or cell propagation (Figure 1A).
Affiliation: Section on Eukaryotic Transposable Elements, Program in Cellular Regulation and Metabolism, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA.