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Primer probe group for detecting rs1045642 and application of primer probe group

A primer-probe and probe technology, applied in recombinant DNA technology, microbial assay/inspection, biochemical equipment and methods, etc., can solve problems such as resistance and drug concentration reduction, and achieve simple operation and great application and promotion value. Effect

Inactive Publication Date: 2018-05-22
山东德诺生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Individuals carrying mutant alleles have changes in the activity of P-glycoprotein, resulting in changes in the transmembrane transport process of various drugs mediated by it in vivo, which makes the abnormal transfer of drugs from the inside of the cell to the outside of the cell against the concentration gradient. The drug concentration inside the cell decreases, leading to drug resistance

Method used

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  • Primer probe group for detecting rs1045642 and application of primer probe group
  • Primer probe group for detecting rs1045642 and application of primer probe group
  • Primer probe group for detecting rs1045642 and application of primer probe group

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Embodiment 1, be used to detect the design of the primer probe set of rs1045642

[0051] After a lot of design, pre-experiment, and effect comparison, a set of primer probes for detecting rs1045642 was obtained.

[0052] Primer F: 5'-TTGCTGAGAACATTGCCTATGGAGA-3';

[0053] Primer R: 5'-TGACTCGATGAAGGCATGTATGTTG-3';

[0054] Probe P: 5'-FAM- ATC GTGAGGGCAGCAAAG GAT -BHQ1-3'.

[0055] Primer F is a single-stranded DNA molecule shown in Sequence 1 of the Sequence Listing. Primer R is a single-stranded DNA molecule shown in Sequence 2 of the Sequence Listing. Probe P is a single-stranded DNA molecule with a FAM fluorescent group at the 5' end and a BHQ1 fluorescent quencher group at the 3' end. The nucleotide sequence of the DNA molecule is shown in sequence 3 in the sequence table, and the underlined nucleosides The acids (ie, positions 1, 2, 3, 19, 20, and 21) are locked nucleic acids (LNA).

[0056] The target sequences of primer F and primer R are shown in seque...

Embodiment 2

[0057] Embodiment 2, the application of the primer probe set for detecting rs1045642

[0058] 1. Blood sample processing (processing method of a single blood sample)

[0059] 1. Take 65 μL of EDTA anticoagulant blood, add 500 μL of lysate, mix by inversion, centrifuge at 12000 rpm for 4 min, and discard the supernatant.

[0060] Lysis solution: 1.7g / 100ml NaCl aqueous solution.

[0061] 2. After completing step 1, add 120 μL of preservation solution, first bathe in water at 65°C for 10 minutes, then in water bath at 85°C for 10 minutes, then centrifuge at 12,000 rpm for 4 minutes, then store at 4°C for later use, and take the supernatant before use, which is the template sample.

[0062] The preparation method of the preservation solution: ① mix 25 parts by volume of FG buffer solution and 4 parts by volume of proteinase K solution to obtain a mixed solution; ② mix 1 part by volume of the mixed solution obtained in step ① with 5 parts by volume of ddH 2 O mixed to obtain a p...

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Abstract

The invention discloses a primer probe group for detecting rs1045642 and application of the primer probe group. The invention firstly protects the primer probe group which comprises a primer F, a primer R and a probe P; the primer F is a single-strand DNA molecule shown as a sequence 1 in a sequence table; the primer R is a single-strand DNA molecule shown as a sequence 2 in the sequence table; the probe P is a single-strand DNA molecule with the one terminus having a fluorescent group and the other terminus having a fluorescence quenching group; a part of nucleotides in the DNA molecule are locked nucleic acids; and a nucleotide sequence of the DNA molecule is shown as a sequence 3 in the sequence table. The primer probe group provided by the invention can be used for detecting rs1045642and judging a genotype of a sample to be detected based on the site so as to guide medication and having an important application and popularization value.

Description

technical field [0001] The invention relates to a primer probe set for detecting rs1045642 and application thereof. Background technique [0002] ATP-binding cassette transporters (ABC transporters) transport a variety of molecules to and from the cell membrane and are divided into seven distinct subfamilies (ABC1, MDR / TAP, MRP, ALD, OABP, GCN20, White). Members of the MDR / TAP family are involved in the multidrug resistance process. P-glycoprotein (P-gp) is a member of the MDR / TAP family. P-glycoprotein is an ATP-dependent drug efflux pump responsible for reducing drug accumulation in drug-resistant cells and mediating the formation of drug resistance to anticancer drugs. P-glycoprotein plays an important role in the absorption process of drugs. P-glycoprotein on the surface of intestinal mucosa affects drug absorption and indirectly affects blood drug concentration by pumping drugs back into the intestinal cavity. P-glycoprotein also acts as a transporter in the blood-br...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12Q1/6883C12Q1/686C12N15/11
CPCC12Q1/686C12Q1/6883C12Q1/6886C12Q2600/156C12Q2563/107C12Q2545/114C12Q2527/107C12Q2525/10
Inventor 丛茜赵海文刘婷婷李朋孙文文杨银辉
Owner 山东德诺生物科技有限公司
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