Targeted Sequencing of Telomere Genes

DNA was extracted from peripheral blood (PureGene Blood Core Kit; Qiagen). Mutations were detected by either PCR amplification and Sanger sequencing, as described previously (11 (link), 41 (link)), whole-exome sequencing (29 (link), 37 (link)), whole-genome sequencing (35 ), or custom amplicon sequencing followed by confirmation with Sanger sequencing (12 (link)). For targeted panel sequencing, we designed a TruSeq Custom Amplicon probe set (Illumina) that included the coding and flanking intronic sequences of telomere genes (TERT, TR, DKC1, RTEL1, NAF1, TINF2, CTC1, PARN, NOP10, NHP2, TCAB1) and GATA2 (including the intron 4 enhancer). Libraries were generated from 500 ng of DNA according to manufacturer recommendations and analyzed on a MiSeq sequencer (Illumina) (12 (link)). The mean coverage of target sequence was 238× and 88% of the target sequences were covered at or greater than 8×.

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Alder J.K., Hanumanthu V.S., Strong M.A., DeZern A.E., Stanley S.E., Takemoto C.M., Danilova L., Applegate C.D., Bolton S.G., Mohr D.W., Brodsky R.A., Casella J.F., Greider C.W., Jackson J.B, & Armanios M. (2018). Diagnostic utility of telomere length testing in a hospital-based setting. Proceedings of the National Academy of Sciences of the United States of America, 115(10), E2358-E2365.

Publication 2018

PureGene Blood Core Kit MiSeq sequencer

Blood Ctc1 Gata2 Genes Intron Mutations Nhp2 Nop10 Rtel1 Telomere Tert Tinf2

Corresponding Organization :

Other organizations : Johns Hopkins University, Johns Hopkins Medicine, Sidney Kimmel Comprehensive Cancer Center

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