Method for detecting the chimerism rate of recipients after allogeneic hematopoietic stem cell transplantation

Abstract

The invention adopts the real-time quantitative PCR (RQ-PCR) technology combining with single nucleotide polymorphism (SNP) to detect the chimerism rate of recipients after allogeneic hematopoietic stem cell transplantation, and establishes a standard curve of a plasmid standard which contains target genes and internal reference and is subjected to multiple proportions dilution. A standard construction template is a qualitative PCR product for a patient with positive SNP loci, the standard construction template is cloned in a T carrier to construct a plasmid, and the standard curve is established after multiple proportions dilution. The new method facilitates early detection and exclusion as well as relapse prevention, improves the success rate of transplantation, has higher sensitivity and stability, does not need to consume a large number of valuable specimens before transplantation, and can be used for individual identification purposes. The invention can replace the present prevailing STP-PCR semi-quantitative detection method, can carry out quantitative detection of the chimerism rate by utilizing real-time quantitative PCR instruments purchased by most hospitals, and has accuracy, simplicity, speediness and low cost.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and more specifically relates to a novel method for detecting the chimerism rate of donors and recipients after allogeneic hematopoietic stem cell transplantation. Background technique [0002] In recent years, the incidence of malignant hematological diseases in children and adults has been increasing year by year. With conventional treatments such as radiotherapy and chemotherapy, the 3-year survival rate is only 40-60%, and it is difficult to be completely cured. Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the only way to achieve a cure. At present, the number of transplant cases at home and abroad is increasing year by year, and dozens of hospitals in China have carried out hematopoietic stem cell transplantation. However, the rejection and recurrence after allo-HSCT have greatly affected the success rate of transplantation. The success of transplantation is ...

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Problems solved by technology

[0006] The article "Quantitative assessment of hematopoieticchimerism after bone marrow transplantation by real-time quantitative polymerase chain reaction" in the 99th issue of "Blood" magazine in 2002 reported the method of RQ-PCR combined with SNP to detect the chimerism rate after transplantation, which is significantly higher than STR-PCR. The sensitivity and accuracy of the detection are improved, but the donor-recipient cells need to be used to simulate donor-recipient chimerism for multiple dilution to establish a standard curve. The standard curve established by this method requires the use of a large number of precious samples before transplantation, and the operation is cumbersome. Increased detection cost and time, and reduced the stability of the experiment

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