Application of product for detecting ddcfDNA in preparing product for detecting transplanted kidney injuries caused by pulmonary infection and evaluating prognosis effect

A technology for lung infection and kidney injury, applied in the field of medical detection, can solve the problems of difficulty in evaluating the effect of lung infection on transplanted kidney injury, inability to effectively detect transplanted kidney injury and prognosis, etc. Painful effects of patients

Pending Publication Date: 2019-11-15
上海奥根诊断技术有限公司
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0006] Therefore, the problem to be solved by the present invention is to overcome the inability to effectively detect the injury and prognosis of the transplanted kidney caused by pulmonary infection in existing clinics, and it is difficult to evaluate the damage of pulmonary infection to the transplanted kidney and the effect after treatment of pulmonary

Method used

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  • Application of product for detecting ddcfDNA in preparing product for detecting transplanted kidney injuries caused by pulmonary infection and evaluating prognosis effect
  • Application of product for detecting ddcfDNA in preparing product for detecting transplanted kidney injuries caused by pulmonary infection and evaluating prognosis effect
  • Application of product for detecting ddcfDNA in preparing product for detecting transplanted kidney injuries caused by pulmonary infection and evaluating prognosis effect

Examples

Experimental program
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Example Embodiment

[0069] Example 1 SNP site screening

[0070] Select the dbSNP (data base SNP, https: / / www.ncbi.nlm.nih.gov / snp / ) database and the Chinese population database (stored in Shanghai Aogen Diagnostics Technology Co., Ltd.). The Chinese population database is based on 3000 cases of Chinese The list of genetic polymorphism loci constructed from the whole genome sample information, such as figure 1 As shown, there is some difference frequency information between the Chinese population database and the Hapmap database (http: / / www.hapmap.org), which represents the different genetic loci in the Chinese population, so that the Chinese population database can more accurately present the Chinese population Information about the genetic polymorphism locus.

[0071] From the above two databases, the sites with a minor allele frequency (MAF) close to 0.5 were selected, and 5754 target SNP sites as shown in Table 1 were obtained.

[0072] The SNP sites of the drug metabolism genes, such as the CYP3A4...

Example Embodiment

[0095] Example 2 Extraction of genomic DNA and plasma cfDNA

[0096] 1. Sample collection and separation of blood cells and plasma

[0097] (1) Sample collection: Divide the blood from the donor into two parts, each about 8ml, save and transport with streck preservation tube;

[0098] (2) Plasma separation: Centrifuge a fresh whole blood sample at 1900g at 4°C for 10 minutes, separate the supernatant into a new centrifuge tube, continue to centrifuge the supernatant at 16000g, at 4°C for 10 minutes, and centrifuge the supernatant Transfer the liquid to a new centrifuge tube to obtain separated plasma, which can be stored at -20°C;

[0099] (3) Blood cell separation: the lower layer of sediment after centrifuging another fresh whole blood sample is the blood cell.

[0100] 2. Use (Laifeng Genomic DNA Extraction Kit, DK601-02) to extract the separated blood cells to obtain genomic DNA; use (QIAamp Circulating Nucleic Acid Kit, 55114) to extract the separated plasma to obtain plasma cfDNA...

Example Embodiment

[0107] Example 3 Library construction

[0108] 1. Take 1μl of plasma cfDNA obtained in Example 2 and perform QuantiFluor TM -ST (Promega) quantification, another 1μl was used to test the quality using Agilent 2100.

[0109] 2. Using the plasma cfDNA and blood cell genomic DNA obtained in Example 2 as samples, use VAHTSUniversal DNA Library Prep Kit for Illumina V3 to prepare plasma cfDNA and genomic DNA libraries. The specific steps are as follows:

[0110] (1) End filling

[0111] a. Add 7.5μl End Repair mix to the labeled centrifuge tube;

[0112] b. Take 25μl of the quantified sample in step 2 and add the end-filling mixture shown in a to obtain a sample reaction solution with a total volume of 32.5μl, and pipette to mix evenly;

[0113] c. Run the following program on the PCR machine:

[0114] Keep at 20℃ for 15min,

[0115] Keep at 65℃ for 15min,

[0116] Let stand at 4°C.

[0117] (2) Add single-molecule labeling linker

[0118] a. Calculate the amount of reagents required according to...

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Abstract

The invention provides application of a product for detecting ddcfDNA in preparing a product for detecting transplanted kidney injuries caused by pulmonary infection and evaluating a prognosis effect,the product for detecting the transplanted kidney injuries caused by pulmonary infection and evaluating the prognosis effect, and a method for detecting the transplanted kidney injuries caused by pulmonary infection and evaluating the prognosis effect. The product for detecting the ddcfDNA can detect the level of the donor free cfDNA (ddcfDNA) in plasma of a kidney transplant recipient, accordingto the level of the donor free cfDNA (ddcfDNA), the transplanted kidney function injuries caused by pulmonary infection are diagnosed, the prognosis effect is evaluated, the early diagnosis of the transplanted kidney injures caused by pulmonary infection and the prognosis effect for the organ transplant recipient after an operation can be achieved, the product for detecting the ddcfDNA has the advantages of rapidity, accuracy and high sensitivity, the detection method is non-invasive, and the pain of the patient is greatly reduced.

Description

technical field [0001] The invention relates to the technical field of medical detection, in particular to the use of a product for detecting ddcfDNA in the preparation of a product for detecting transplanted kidney injury and prognosis caused by lung infection, for detecting the effect of lung infection on transplanted kidney injury and prognosis products, and methods for detecting the impact of pulmonary infection on transplanted kidney injury and prognosis. Background technique [0002] Infection is one of the most common complications after organ transplantation. Although with the development and use of new immunosuppressive drugs, which can reduce the incidence of rejection and prevent and treat infectious complications, the survival rate of organ transplant recipients has increased significantly in the past few decades. However, infection is still an important cause of organ transplant failure. [0003] Immunosuppressants are used in patients after renal transplantat...

Claims

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Application Information

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IPC IPC(8): C12Q1/6883C40B50/06C12Q1/6869
CPCC12Q1/6883C40B50/06C12Q1/6869C12Q2600/118C12Q2600/156C12Q2535/122
Inventor 蒋廷亚刘海涛葛军刘枫王啸晨黎小锋
Owner 上海奥根诊断技术有限公司
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