Method for detecting rearrangement clonality of correlative genes of lymphocyte

Abstract

The invention discloses a method for detecting rearrangement clonality of correlative genes of lymphocyte and belongs to the technical field of biology. According to the method, a high-throughput next-generation sequencing method is adopted for detecting rearrangement clonality of the correlative genes of lymphocyte tissue, and specifically, a multifunctional primer and a PCR reagent are utilizedto conduct multiplication on samples to be detected at first to obtain a multi-sample gene locus multiplication sublibrary, wherein both ends of the multiplication sublibrary are connected with DNA segments of different connectors, one side of the multiplication sublibrary is a sequencing connector which can contain a sample tag for distinguishing sequencing results of the different samples, and the other side of the multiplication sublibrary is a fixed connector used for connecting caught particles; then, high-throughput sequencing is conducted on the multiplication sublibrary, sequencing results are subjected to classified sorting according to a tag sequence, the samples and target genetic loci, and whether or not there is clonality rearrangement of target genes is analyzed. The method has the advantages that the detection throughput, the sensitivity and the specificity are high, the sensitivity can reach 0.01%, and the DNA concentration of the samples can be lowered to 0.1-1ng / microliter.

Description

technical field [0001] The invention relates to a method for detecting the clonality of lymphocyte-related gene rearrangement, and belongs to the field of biotechnology. Background technique [0002] Lymphocytes are produced by lymphoid organs and are a type of cell line with the body's immune recognition response function, which can be divided into B cells, T cells and NK cells. There are receptor proteins on the surface of lymphocytes that can participate in immune reactions, and can specifically bind to antigens or antibodies, such as immunoglobulin (IG) on the surface of B lymphocytes and T cell receptor (TCR) on the surface of T lymphocytes. Taking the IGH gene expressing immunoglobulin heavy chain polypeptide as an example, in undifferentiated lymphocytes, the IGH gene is mainly composed of a variable region (variabl V), a diversity region (diversity D), and a joining region (joining J). Each gene region is composed of multiple gene segments. With the differentiation...

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Problems solved by technology

However, there are still big problems in the clinical application of this method of detection: 1. There are many gene sites to be detected, and one sample needs multiple PCR reactions for detection, which increases the detection cost and complicates the operation process; 2. It cannot be performed More detailed and in-depth pedigree analysis can only detect monoclonal rearrangements for IG or TCR genes, but cannot in-depth analysis of which family gene in the V region or J region is involved in the rearrangement, providing more detailed information for the clinic; 3. Detection limit There are still major limitations. The current PCR-capillary electrophoresis method can only detect 1% of monoclonal rearrangements, which cannot meet the needs of clinical MRD detection; 4. Can not detect somatic hypermutation at the same time

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