b'Next-Generation Sequencing (NGS) | Research Use Only (RUO) AssaysIGK Assay Next-Generation Sequencing (NGS) | RUO AssaysLymphoTrack IGK AssayAssay Uses Background This research use only (RUO) assay for next-generation sequencingThe human immunoglobulin kappa (IGK) gene locus on chromosome (NGS), identifies clonal IGK V-J, V-K de , and intron-K de(INTR-K de )2 (2p11.2) includes 76 V (variable) region genes spanning 7 subgroups rearrangements, the corresponding DNA sequences, and providesand 5 J (joining) region gene segments upstream of the C region. the distribution frequency of V-J, V-K de , and INTR-K desegmentThe K de , approximately 24 kb downstream of the J-C region, can also utilization using the Illumina MiSeq or Thermo Fisher Scientificrearrange with V gene segments and the isolated recombination Ion PGM and Ion S5 platforms. signal sequence in the J-C intronic region.During development of lymphoid cells, antigen receptor genes Summary and Explanation of the Testundergo somatic gene rearrangements. Specifically during B-cell development, genes encoding IGK molecules are assembled from The LymphoTrack IGK Assay represents a significant improvementmultiple polymorphic gene segments that undergo rearrangements over clonality assays that utilize fragment analysis by providing threegenerating V-J combinations unique in both length and sequence. Since important and complementary uses: leukemias and lymphomas originate from the malignant transformation 1.Detection of clonal populations. of individual lymphoid cells, all leukemias and lymphomas generally share one or more cell-specific or clonal antigen receptor gene 2. Identification of sequence information and gene rearrangements. Therefore, tests that detect IGK clonal populationssegment utilization. can be useful in the study of B-cell malignancies and complement IGH 3. Ability to track sequences in subsequent samples with the testing, as the IGK receptor is less susceptible to somatic mutations. use of the LymphoTrack MRD Software.Unlike conventional fragment analysis assays, this NGS methodSpecimen Requirementutilizes a single multiplex master mix to target conserved regions50 ng of high-quality genomic DNA.of IGK that are described in lymphoid malignancies. Primers are designed with Illumina adapters and indices (8-24) or Thermo Fisher Scientific adaptors and indices (12), thereby allowing fora one-step PCR reaction to generate sequence-ready amplicons. ReferencesTo reduce per sample testing costs, amplicons from different1.S Tonegawa et al., Nature 302: 575-581 (1983).samples (amplified with different indexed master mixes) or 2. JE Miller et al., Molecular Genetic Pathology (2nd ed.). Springer LymphoTrack kits can be sequenced together on a single IlluminaMiSeq flow cell, Ion S5 or PGM chips. Science & Business Media. 2013: 30.2.7.13 and 30.2.7.18.The LymphoTrack bioinformatics software enables simplified3. KJ Trainor et al., Blood 75: 2220-2222 (1990).analysis and visualization of data and the LymphoTrack MRD Software allows identified 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 the IGK GeneDepicted are the variable region (V) genes or variable intragenic J-C intron (J-C INTR) and downstream consensus joining region genes (J) or kappa deleting Kdeelement (K de ) that are involved in IGK gene rearrangements.Kde 42'