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Reagent kit for detecting agedness yellow spot degenerative disease

A technology for degenerative diseases and age-related macular, which can be used in measurement devices, microbial determination/inspection, instruments, etc., and can solve problems such as poor results and difficult vision functions

Active Publication Date: 2011-08-31
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

[0040] [Example 1] Blood sample collection and specimen processing

[0041] A total of 214 Chinese patients with age-related macular degeneration (AMD) and 214 controls were selected. The patients were clinically confirmed AMD, including 115 patients with wet AMD and 99 patients with dry AMD. The controls were people who were older than or equal to 60 years old, had no drusen and retinal pigment changes, and matched the age group of AMD patients.

[0042] For all participants, a detailed investigation of their medical and family histories, and a physical examination and detailed ophthalmology specialist examination were performed. After signing the informed consent form, 5-10 ml of EDTA anticoagulated blood samples were collected from each person. Genomic DNA was extracted by the phenol-chloroform method.

Embodiment 2

[0043] [Example 2] PCR amplification and genotype analysis of the deletion / insertion variant gene at 4340 bases upstream of the HTRA1 gene

[0044] (1) Electrophoretic separation and identification of PCR amplification products

[0045] 1. Take the blood of the subject and extract the DNA;

[0046] 2. With 5'-GCCAACTGGAGCTTCTCATC-3' and

[0047] 5'-GTAGTTTCCAGGGGCTCTCC-3' is the upstream and downstream primers for PCR amplification of genomic DNA; reaction conditions:

[0048] 95℃3minutes, (94℃30seconds, 57℃30seconds, 72℃45seconds)*35cycles

[0049] 3. Separation by 1% agarose gel electrophoresis, and discrimination of deletion / insertion variation based on the size of the PCR product.

[0050] Agarose electrophoresis identification:

[0051] The PCR product containing the deletion / insertion sequence of 4340bp 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, an...

Embodiment 3

[0086] [Example 3] PCR amplification and genotype analysis of DNA fragments containing rs2293870, rs11200638, and rs10490924 single nucleotide polymorphisms

[0087] 1. Extract the blood genome DNA of AMD patients and control groups.

[0088] 2. PCR amplification of genomic DNA fragments containing target sequences (rs2293870, rs11200638, rs10490924)

[0089] Primer sequence:

[0090] rs2293870: Forward: 5'-GGCCGCTCGGCGCCTTTGGC-3'

[0091] Reverse: 5'-CCCCGAAGGGCACCACGCAC-3'

[0092] rs11200638: Forward: 5'-ATGCCACCCACAACAACTTT-3'

[0093] Reverse: 5'-CGCGTCCTTCAAACTAATGG-3'

[0094] rs10490924: Forward: 5'-TACCCAGGACCGATGGTAAC-3';

[0095] Reverse: 5'-GAGGAAGGCTGAATTGCCTA-3'

[0096] Reaction conditions:

[0097] rs2293870: 95℃3minutes, (94℃30seconds, 57℃30seconds, 72℃45seconds)*35cycles

[0098] rs11200638: 95℃3minutes, (94℃30seconds, 52℃30seconds, 72℃45seconds)*35cycles

[0099] rs10490924: 95℃3minutes, (94℃30seconds, 55℃30seconds, 72℃45seconds)*35cycles

[0100] ...

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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 highassociation 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 disease, in particular to detecting 443 bases in the part of 3'UTR region of LOC387715 gene at 4340 bp upstream of HTRA1 gene which is related to age-related macular degeneration disease Base sequence deletion / 54 base sequence insertion variation kit. Background technique [0002] Macular degeneration is considered to be a heterogeneous disease characterized by progressive central vision loss and macular retinal pigment epithelial degeneration, and is an important blinding eye disease. Macular degeneration includes age-related macular degeneration and macular dystrophy (stargart disease, best disease, sorsby fundus dystrophy, etc.). Age-related macular degeneration (AMD) has become one of the most important blinding eye diseases for the elderly, with 10 million patients worldwide, and the prevalence of AMD among people aged 60-69 in my country is 7.77%. Over the age ...

Claims

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Application Information

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Patent Type & Authority Patents(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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