Sequencing-Based Fusion Detection

DNA and RNA were co-isolated from each specimen as previously described.(23 ) DNA and RNA libraries were generated per sample using the Ion Ampliseq Library kit (Life Technologies, Foster City, CA), as described.(23 ) We prepared templates for DNA and RNA libraries using the Ion PI Template OT2 200 Kit v3 on the Ion One Touch 2 and sequencing was performed on Ion Proton P1 chips using the Ion PI Sequencing 200 Kit v3 (200 base pair reads), essentially as described.(24 (link),25 ) NGS data analysis was performed using Torrent Suite (4.2.0) and the Coverage Analysis Plug-ins (both v4.0-r73765), along with the Ion Reporter (4.2.0) Targeted NGS, fusion analysis workflow and in house validated pipelines as described in the Supplementary Methods.(24 (link)-27 ) A sample was classified as fusion positive if a fusion isoform was supported by ≥ 20 reads and ≥ 3.0% total mapped reads, otherwise is classified as fusion negative.

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Palapattu G.S., Salami S.S., Cani A.K., Hovelson D.H., de la Vega L.L., Vandenberg K.R., Bratley J.V., Liu C.J., Kunju L.P., Montgomery J.S., Morgan T.M., Natarajan S., Huang J., Tomlins S.A, & Marks L.S. (2016). Molecular Profiling to Determine Clonality of Serial Magnetic Resonance Imaging/Ultrasound Fusion Biopsies from Men on Active Surveillance for Low Risk Prostate Cancer. Clinical cancer research : an official journal of the American Association for Cancer Research, 23(4), 985-991.

Publication 2016

Ion Ampliseq Library kit Ion PI Template OT2 200 Kit v3 Ion PI Sequencing 200 Kit v3

Base pair Chips Isoform Library Pi 200 Proton Touch

Corresponding Organization : University of California, Los Angeles

Other organizations : Duke University

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Variable analysis

independent variables

DNA and RNA library generation using the Ion Ampliseq Library kit
Template preparation for DNA and RNA libraries using the Ion PI Template OT2 200 Kit v3
Sequencing on Ion Proton P1 chips using the Ion PI Sequencing 200 Kit v3

dependent variables

Fusion isoform supported by ≥ 20 reads and ≥ 3.0% total mapped reads

control variables

DNA and RNA co-isolation from each specimen as previously described

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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