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Reagent kit for detecting agedness yellow spot degenerative disease

A technology for degenerative diseases and senile macula, applied in measuring devices, microbiological determination/inspection, instruments, etc., can solve problems such as poor results and difficult vision

Active Publication Date: 2009-10-07
SICHUAN ACADEMY OF MEDICAL SCI SICHUAN PROVINCIAL PEOPLES HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Once wet AMD develops, the course of the disease is rapid, the effect of conventional treatment is not good, and it is difficult to restore the lost visual function to a large extent

Method used

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  • Reagent kit for detecting agedness yellow spot degenerative disease
  • Reagent kit for detecting agedness yellow spot degenerative disease
  • Reagent kit for detecting agedness yellow spot degenerative disease

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] [Example 1] Blood sample collection and specimen processing

[0040] 214 Chinese patients with age-related macular degeneration (AMD) and 214 controls were selected. The patients were clinically diagnosed AMD, including 115 wet AMD patients and 99 dry AMD patients. The control group was a group of AMD patients who were older than or equal to 60 years old and had no drusen and retinal pigment changes.

[0041] A detailed investigation of the medical and family history of all participants, physical examination and detailed ophthalmological examinations were performed. After signing the informed consent form, each person collects 5-10ml of EDTA anticoagulant blood sample. The phenol-chloroform method was used to extract genomic DNA.

Embodiment 2

[0042] [Example 2] PCR amplification and genotype analysis of the 4340 base upstream deletion / insertion variant gene containing HTRA1 gene

[0043] (1) Electrophoresis separation and identification of PCR amplification products

[0044] 1. Take blood from the subject and extract DNA;

[0045] 2. Using 5’-GCCAACTGGAGCTTCTCATC-3’ and 5’-GTAGTTTCCAGGGGCTCTCC-3’ as upstream and downstream primers, PCR amplifies genomic DNA;

[0046] Reaction conditions:

[0047] 95℃3minutes, (94℃30seconds, 57℃30seconds, 72℃45seconds)*35cycles

[0048] 3. 1% agarose gel electrophoresis separation, based on the size of PCR product to distinguish deletion / insertion variation.

[0049] Agarose electrophoresis identification:

[0050] The PCR product containing the 4340bp deletion / insertion sequence upstream of the HTRA1 gene was separated by 1% agarose gel electrophoresis. The deletion / insertion made the normal fragment of the PCR product about 400bp smaller, and the genotype could be judged by electropho...

Embodiment 3

[0070] [Example 3] PCR amplification and genotype analysis of DNA fragments containing rs2293870, rs11200638, and rs10490924 SNPs

[0071] 1. Extract blood genomic DNA from AMD patients and control groups.

[0072] 2. PCR amplification of genomic DNA fragments containing the target sequence (rs2293870, rs11200638, rs10490924)

[0073] Primer sequence:

[0074] rs2293870: Forward: 5’-GGCCGCTCGGCGCCTTTGGC-3’

[0075] Reverse: 5’-CCCCGAAGGGCACCACGCAC-3’

[0076] rs11200638: Forward: 5’-ATGCCACCCACAACAACTTT-3’

[0077] Reverse: 5’-CGCGTCCTTCAAACTAATGG-3’

[0078] rs10490924: Forward: 5’-TACCCAGGACCGATGGTAAC-3’;

[0079] Reverse: 5’-GAGGAAGGCTGAATTGCCTA-3’

[0080] Reaction conditions:

[0081] rs2293870: 95℃3minutes, (94℃30seconds, 57℃30seconds, 72℃45seconds)*35cycles

[0082] rs11200638: 95℃3minutes, (94℃30seconds, 52℃30seconds, 72℃45seconds)*35cycles

[0083] rs10490924: 95℃3minutes, (94℃30seconds, 55℃30seconds, 72℃45seconds)*35cycles

[0084] 3. Geno...

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Abstract

The present invention provides a reagent kit for detecting an agedness yellow spot degenerative disease, capable of being used to early monitoring and detecting AMD high risk groups, so as to prevent and treat early to reduce eyesight injury. The reagent kit takes a deletion / insertion variation (locating at a 4340 base point of the HTRA1 gene upstream, containing 443 base sequence deletion of LOC387715 gene part 3'UTR region and 54 base sequence inserted variation) between LOC387715 gene and HTRA1 gene, a HTRA1 gene no.1 exon G-> mutation and relevant linkage sites / genes as target, has a high association degree with the AMD, and further improves the detecting accuracy by cooperating with the analysis of HTRA1 gene promoter region G->A variation (mononucleotide polymorphism SNP rs11200638) and the HTRA1 gene upstream sequence G->T variation (mononucleotide polymorphism SNP rs10490924).

Description

Technical field [0001] The present invention relates to a kit for detecting age-related macular degeneration diseases, specifically detecting the 443 bases between LOC387715 gene and HTRA1 gene related to age-related macular degeneration disease, 4340bp upstream of HTRA1 gene containing part of the 3'UTR region of LOC387715 gene Base sequence deletion / 54 base sequence insertion mutation kit. Background technique [0002] Macular degeneration is considered to be a heterogeneous disease characterized by progressive central vision decline and macular retinal pigment epithelial degeneration, and is an important blind eye disease. Macular degeneration includes age-related macular degeneration, macular dystrophy (stargart disease, best disease, sorsby ocular dystrophy, etc.). Age-related macular degeneration (AMD) has become one of the most important blinding eye diseases in the elderly. There are 10 million patients worldwide. The prevalence of AMD in the 60-69-year-old population in ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N33/573
Inventor 杨正林张康鲁芳林婴
Owner SICHUAN ACADEMY OF MEDICAL SCI SICHUAN PROVINCIAL PEOPLES HOSPITAL
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