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. 1These developmentally regulated gene The LymphoTrack TRG Assay represents a significant improvementrearrangements generate V-J combinations that are unique for over existing clonality assays that utilize fragment analysis byeach cell. 2Since leukemias and lymphomas originate from the providing three important and complementary uses: malignant transformation of individual lymphoid cells, all leukemias 1.Detection of clonal populations. and 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 This assay utilizes a single multiplex master mix to target conservedclonal / as well as clonal / T-cells. Clonal / T-cells would be V and J regions of the human TRB gene locus described in lymphoidexpected to have biallelic TRG gene rearrangements. malignancies, thereby reducing sample DNA requirements and simplifying the testing workflow. Primers are designed with Illumina Note: For a more thorough explanation of the locus and the targeted deep sequencing strategy, please refer to Principle of Immunoglobulin adapters and indices (8-24) or Thermo Fisher Scientific adaptorsand T-Cell Receptor Gene Rearrangement. 2and indices (12), thereby allowing for a one-step PCR reaction to generate sequence-ready amplicons. In addition, amplicons from different samples (amplified with different indexed master mixes) orSpecimen RequirementLymphoTrack kits can be sequenced together on a single IlluminaMiSeq flow cell Ion S5 or PGM chip to reduce per sample testing50 ng of high-quality genomic DNA.costs.The LymphoTrack bioinformatics software enables simplifiedReferencesanalysis and visualization of data generated from this assay and1.Tonegawa, S. (1983). Nature 302, 575-581.the LymphoTrack MRD Software allows sequences to be tracked in2. Miller JE. (2013) Molecular Genetic Pathologysubsequent samples. Please see the LymphoTrack MRD software(2nd Edition.,sections 30.2.7.13 and 30.2.7.18).section to learn how the LymphoTrack Assays can be applied to MRD studies, or email [email protected] (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.50'