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. 2019 Aug 30;33(3):334–343. doi: 10.1038/s41379-019-0359-9

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — A representative acute myeloid leukemia case harboring an FLT3-internal tandem duplication tested by fragment analysis and next generation sequencing. a FLT3 mutational analysis by fragment analysis test showing the presence of an FLT3-internal tandem duplication; x axis represent the PCR product size in base pair (bp) and y axis represent the fluorescence intensity. The wild-type peak at 330 bp (black arrow) and FLT3-internal tandem duplication peak at 366 bp (red arrow) were indicated. The FLT3-internal tandem duplication/wild-type allelic ratio was 0.7 as calculated from the internal tandem duplication and wild-type peak fluorescence intensity ratio. b Next generation sequencing read alignments displayed in Alamut® Visual demonstrating the presence of an FLT3-internal tandem duplication (c.1751_1786dup; p.R595_E596ins12) with a variant allele fraction (FLT3-internal tandem duplication/total) of 40% and a corresponding allelic ratio (FLT3-internal tandem duplication/wild type) of 0.7. FLT3-internal tandem duplication reads showed only partial alignment with the reference sequence and 5′ and 3′ soft-clipped bases (colored gray) flanking the duplicated wild-type sequence of the FLT3-internal tandem duplication (blue box). The 3′ soft-clipped bases mark the FLT3-internal tandem duplication insertion site and the FLT3-internal tandem duplication sequences. Red stars mark the wild-type reads with complete alignment with the reference sequence. Green, forward reads; blue, reverse reads. Note that the FLT3 coding reference sequence is in reverse complementary orientation to the next generation sequencing Binary Alignment Map alignment sequence