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Kit for evaluating lamivudine dosage

A technology of lamivudine and kit, which is applied in the fields of molecular biology and medicine, can solve the problems of decreased binding force of triphosphate, uninhibited and lost enzymatic process, etc.

Inactive Publication Date: 2009-02-11
XINBAXIANG SHANGHAI MOLECULAR MEDICAL TECH SHANGHAI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The YMDD part of the wild strain is in a relaxed pocket shape, which is conducive to the combination of nucleotides and lamivudine. The mutation at position 552 makes the side chain shorter and less flexible, and it is between HBV polymerase and lamivudine triphosphate The formation of steric hindrance leads to a decrease in the binding force with lamivudine triphosphate, so that the synthesis of HBV DNA cannot continue
[0006] After YMDD mutation, the lamivudine binding target loses the normal motif of YMDD, and the sensitivity of the drug binding is greatly reduced, so that lamivudine can reverse transcribe the HBV pregene RNA and copy the negative-strand DNA to positive-strand DNA The enzymatic process cannot be inhibited, resulting in drug resistance

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1. Use of detection kits

[0039] Step 1: Extraction of DNA template

[0040] Genomic DNA of oral epithelial cells was extracted by silica gel adsorption.

[0041] Step 2: PCR amplification reaction

[0042] Use the PCR reaction set included in the detection kit.

[0043] The reaction system has a total volume of 25 μl, including 1 μl of DNA template with a concentration of 12.5ng / μl, 2.5 μl of 10X PCR reaction buffer, 0.2 μl of 25mM dNTP mixture, 1.5 μl of 25mM MgCl2 solution, 0.125 μl of 5 units / μl Taq DNA polymerase, 0.25 μl of each primer for 20 μM specific primer pair, 19.175 μl of deionized water.

[0044] The reaction was carried out on the ABI2720 type PCR amplification instrument. The reaction conditions were 94°C, 12 minutes, and 30 cycles of 94°C, 30 seconds, 60°C, 30 seconds, 72°C, 30 seconds, and then 72°C, 10 minutes .

[0045] Step 3: PCR Product Purification

[0046] Use the PCR product purification kit in the detection kit. The reaction sy...

Embodiment 2

[0054] Embodiment 2. The service that carries out lamivudine drug susceptibility assessment to the patient

[0055] Step 1: DNA Extraction

[0056] The physicians in the laboratory department of the hospital instructed the patients to use oral swabs to sample oral epithelial cells, and the silica gel adsorption method was used to extract DNA from oral epithelial cells.

[0057] Step 2: Genotyping Assays

[0058] Using the kit provided by the invention, DNA sequencing is performed on the YMDD region of the hepatitis B virus DNA in the patient's body to determine the genotype of the YMDD region.

[0059] Step 3: Assessment of Individual Lamivudine Sensitivity Analysis

[0060] Through the analysis of the genotype of the YMDD region of the patient's hepatitis B virus DNA, an evaluation and analysis report sheet for individual lamivudine drug sensitivity is issued. The report details the patient's sensitivity to lamivudine, which can be used as a reference for physicians to adjus...

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Abstract

The invention discloses a kit for detecting the sensitivity of an individual patient to Lamivudine drugs. The kit comprises a specific primer pair and DNA sequencing primer for detecting the YMDD region gene type of the DNA of hepatitis B in vivo of a patient, a PCR reaction assembly, a PCR product purification assembly, a DNA sequencing reaction assembly and the like. The kit is used for analyzing and evaluating the sensitivity of the patient to the Lamivudine drugs.

Description

technical field [0001] The invention relates to the fields of molecular biology and medicine, and more specifically, the invention relates to a kit for detecting individual drug sensitivity to lamivudine. Background technique [0002] Hepatitis B virus (hepatitis B for short) is liver inflammatory damage caused by hepatitis B virus (hepatitis B virus for short), and is the most widespread and serious infectious disease in China. The clinical manifestations are fatigue, loss of appetite, nausea, vomiting, oily, diarrhea and abdominal distension. Some cases have fever and jaundice, and about half of the patients have insidious onset. Most hepatitis B can be cured after treatment in the acute phase, and the course of many cases is delayed or becomes chronic, and some of them can develop post-hepatitic cirrhosis or even liver cancer; a very small number of cases develop rapidly, and large necrosis of liver cells occurs. Severe hepatitis; others become asymptomatic carriers of t...

Claims

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Application Information

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IPC IPC(8): G01N33/50C12Q1/68
Inventor 冯哲民邹祖烨
Owner XINBAXIANG SHANGHAI MOLECULAR MEDICAL TECH SHANGHAI
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