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Epidermal growth factor receptor inhibitor curative effect predication reagent kit and its uses

A technology of epidermal growth factor and receptor inhibitors, which is applied in the field of biotechnology and clinical medical testing, can solve the problems of gene modification polymorphism differences among different species, and achieve the effect of high specificity and high detection sensitivity

Inactive Publication Date: 2008-03-26
GUANGDONG GENERAL HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This phenomenon suggests that the polymorphic differences in gene modification among different species may lead to the above phenomenon

Method used

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  • Epidermal growth factor receptor inhibitor curative effect predication reagent kit and its uses

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] 1. Materials:

[0031] Taq DNA polymerase and dNTP were purchased from Promega Company of the United States, sequencing reagents (BigDye Terminator kit 3.1) and gene scanning standard (LIZ-500) were purchased from Applied Biosystems Company of the United States; the 3100 type sequencer was a product of Japan TOSHIBA Company, and the 9600 type PCR The instrument is a product of Perkin Elmer Company in the United States.

[0032] 2. Primer and probe design and synthesis:

[0033] According to the base sequence of EGFR in Genebank (GenBank accession number AF288738) as a template, use Primer5 primer design software (U.S. Perkin Elmer Company), design two pairs of nested amplification primers for amplifying the EGFR Intron1 target fragment:

[0034] First round of primers:

[0035] F: 5'_-GGA-TCG-CGG-GAC-TCT-TGA-3';

[0036] R: 5'_-GTG-GGT-TTA-TGG-TCG-GTA.-3'

[0037] Second round of primers:

[0038] F: 5'_-GGC-TCA-CAG-CAA-ACT-TCT-CC-3';

[0039] R: 5'_-CGG-TAG-TCA-C...

Embodiment 2I

[0062] The detection of embodiment 2Intron 1 (CA) n

[0063] (1), PCR amplification target fragment

[0064] 1. Primers

[0065]According to the base sequence of EGFR in Genebank, use Primer5 primer design software to design two pairs of nested amplification primers for amplifying the target fragment of EGFR Intron1:

[0066] First round of primers:

[0067] F: 5'_-GGA-TCG-CGG-GAC-TCT-TGA-3';

[0068] R: 5'_-GTG-GGT-TTA-TGG-TCG-GTA.-3'

[0069] Reaction system: 50ul system

[0070] Reaction conditions: 95°C, 4min; (94°C for 45s, 50°C for 1min, 72°C for 1min)×35cycle, 72°C for 10min

[0071] Second round of primers:

[0072] F: 5'_-GGC-TCA-CAG-CAA-ACT-TCT-CC-3';

[0073] R: 5'_-CGG-TAG-TCA-CGA-AGC-CAG-A.-3'

[0074] Reaction system: 50ul system

[0075] Reaction conditions: 95°C, 4min; (94°C for 45s, 55°C for 1min, 72°C for 1min)×35cycle, 72°C for 10min

[0076] 2. Purification of amplified fragments: After PCR, run electrophoresis on 5ul products to understand the am...

Embodiment 3

[0098] Embodiment 3, amplified fragment gene scanning

[0099] 1. PCR includes increasing target fragments:

[0100] Primers:

[0101] F: 5'_-GGA-TCG-CGG-GAC-TCT-TGA-3';

[0102] R: 5'_-GTG-GGT-TTA-TGG-TCG-GTA.-3'

[0103] Primers need to be labeled with 5’-Fam fluorescent markers.

[0104] 2. Reaction system: 50ul system

[0105] 3. Reaction conditions: 95°C, 4min; (94°C for 45s, 60°C for 1min, 72°C for 1min)×35cycle, 72°C for 10min

[0106] 4. Purification of amplified fragments: After PCR, run electrophoresis on 5ul of the product to understand the amplification efficiency.

[0107] 5. Denaturation: Take 2 ul of the amplified PCR product, add 10 ul of formamide and standard mixture, heat in a PCR instrument at 95°C for 4 min, place it on ice immediately, and load the sample for scanning.

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Abstract

The invention belongs to the biotechnology and clinical medicine detection field, which discloses a kind of regent box and its application for detecting the Intronl(CA)n of epidermal growth factor receptor(EGFR). It can select the malignancy sufferer who can be cured by the selective micromolecule inhibitor for the EGFR tyrosine kinase.

Description

[technical field] [0001] The invention relates to a kit for detecting and classifying human epidermal growth factor receptor (epidermal growth factor receptor, EGFR) Intronl (CA) n microsatellite polymorphism, which can be used to screen for selective EGFR The invention relates to a small molecule inhibitor of tyrosine kinase for effectively treating malignant tumor patients, and belongs to the field of biotechnology and clinical medicine testing. [Background technique] [0002] In recent years, with the development of molecular biology, molecular genetics and molecular pharmacology, especially genomics, people have gradually realized that the different responses of different individuals to the same drug are mostly due to genetic differences. Therefore, on the basis of pharmacogenetics, pharmacogenomics has been developed. Its main purpose is not to discover human genome genes, but to use known gene theory to improve patient treatment, with the goal of drug effect and safety...

Claims

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Application Information

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IPC IPC(8): G01N21/64G01N33/53C12Q1/68
Inventor 吴一龙聂强郭爱林张绪超林嘉颖
Owner GUANGDONG GENERAL HOSPITAL
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