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The disadvantages of RNA-based NGS are those common to all NGS techniques as well as those specific to the RNA-based NGS.

The disadvantages of RNA-based NGS are summarised in the following table.

Table 24: Disadvantages of RNA-based NGS for Testing for NTRK Gene Fusions


  • Cost (high cost per sample compared with other methods; however, targeted sequencing of a subset of genes or transcripts can reduce costs [1])
  • Turnaround time of approximately 1 to 3 weeks (complexity and labour intensity of testing has limited the widespread inclusion in laboratories [2, 3])
  • Labile nature of RNA: RNA quality in formalin-fixed paraffin-embedded tissue may be too poor for clinical testing, especially in older blocks [4, 5]
  • The amount of data produced places substantial demands on bioinformatics and information technologies at all stages of the protocol [6]


  1. Beadling C, Wald AI, Warrick A et al. A Multiplexed Amplicon Approach for Detecting Gene Fusions by Next-Generation Sequencing. J Mol Diagn 2016; 18: 165-175.
  2. Zehir A, Benayed R, Shah RH et al. Mutational landscape of metastatic cancer revealed from prospective clinical sequencing of 10,000 patients. Nat Med 2017; 23: 703-713.
  3. Chen Y, Chi P. Basket trial of TRK inhibitors demonstrates efficacy in TRK fusion-positive cancers. J Hematol Oncol 2018; 11: 78.
  4. Choi Y, Won YJ, Lee S et al. Cytoplasmic TrkA Expression as a Screen for Detecting NTRK1 Fusions in Colorectal Cancer. Transl Oncol 2018; 11: 764-770.
  5. Murphy DA, Ely HA, Shoemaker R et al. Detecting Gene Rearrangements in Patient Populations Through a 2-Step Diagnostic Test Comprised of Rapid IHC Enrichment Followed by Sensitive Next-Generation Sequencing. Appl Immunohistochem Mol Morphol 2017; 25: 513-523.
  6. Rizzo JM, Buck MJ. Key principles and clinical applications of "next-generation" DNA sequencing. Cancer Prev Res (Phila) 2012; 5: 887-900.

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