b'Next-Generation Sequencing (NGS) | Research Use Only (RUO) AssaysTRG Assay Next-Generation Sequencing (NGS) | RUO AssaysLymphoTrack TRG AssayAssay Uses Background This research use only (RUO) assay for next-generation sequencingThe human T-cell receptor gamma (TRG) gene locus on chromosome (NGS) identifies clonal TRG V-J rearrangements and the associated7 (7q14) includes 14 variable region (Group I, II, III, and IV), 5 joining V-J region DNA sequences. It also provides the frequency distributionregion (J) gene segments, and 2 constant (C) genes spread overof V-J segment utilization using the Illumina MiSeq or Thermo Fisher200 kilobases. Scientific Ion S5 and PGM instruments.During development of lymphoid cells, the antigen receptor genes Summary and Explanation of the Test(including gene segments within the TRG locus), undergo somatic gene rearrangement. These developmentally regulated gene The LymphoTrack TRG Assay represents a significant improvementrearrangements generate V-J combinations that are unique for each over existing clonality assays that utilize fragment analysis bycell. Since leukemias and lymphomas originate from the malignant providing three important and complementary uses: transformation of individual lymphoid cells, all leukemias and 1.Detection of clonal populations. lymphomas generally share one or more cell-specific or clonal antigen receptor gene rearrangements. Therefore, tests that detect 2. Identification of sequence information and geneTRG clonal populations can be useful in the study of T-cell and certain segment utilization. B-cell malignancies.Since the TRG locus rearranges before the TRB 3. Ability to track sequences in subsequent samples with the use oflocus, and all unsuccessful or unproductive rearrangements arethe Invivoscribe MRD Software. retained in the cells, examination of the TRG locus can identify both The single multiplex master mix targets all conserved regions withinclonal / as well as clonal / T-cells. Clonal / T-cells would be the variable (V) and the joining (J) genes described in lymphoidexpected to have biallelic TRG gene rearrangements.malignancies. The average PCR amplicon size of the TRG gene rearrangements generated using this assay was designed to beSpecimen Requirementcompatible with fragmented DNA isolated from challenging samples50 ng of high-quality genomic DNA.such as FFPE sections.Primers are designed with Illumina adapters and indices (8-24) or Thermo Fisher Scientific adaptors and indices (12), thereby allowingReferencesfor a one-step PCR reaction to generate sequence-ready amplicons.1.S Tonegawa et al., Nature 302: 575-581 (1983).In addition, amplicons from different samples (amplified with different indexed master mixes) or LymphoTrack kits can be sequenced together2. JE Miller et al., Molecular Genetic Pathology (2nd ed.). Springeron a single Illumina MiSeq flow cell Ion S5 or PGM chip to reduceScience & Business Media. 2013: 30.2.7.13 and 30.2.7.18.per sample testing costs. 3. KJ Trainor et al., Blood 75: 2220-2222 (1990).The LymphoTrack bioinformatics software enables simplified analysis and visualization of data generated from this assay and the LymphoTrack MRD Software allows sequences to be tracked in subsequent samples. Please see the LymphoTrack MRD software section to learn how the LymphoTrack Assays can be applied to MRD studies, or email marketing@invivoscribe.com. Positive (clonal) and negative (polyclonal) DNA controls are included in the kits. Simplified Representation of theTRG GeneSimple representation of the organization of the T-cell receptor gamma gene on chromosome 7. Depicted are the variable region genes (V2V11) and downstream joining region genes (J1/2JP1/2) that are involved in rearrangements in T-cell lymphomas.44'