Method for screening HRDs disease-causing mutation and gene chip hybridization probe designing method involved in same

A gene chip hybridization and hybridization probe technology, applied in the field of biomedicine, can solve the problems of screening out disease-causing genes, not being able to locate disease-causing sites, and being difficult to analyze small families and sporadic cases, etc., to achieve high screening efficiency, screening Check the effect of a wide range of

Active Publication Date: 2014-03-26
赵晨
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AI Technical Summary

Problems solved by technology

The positional cloning strategy based on linkage analysis is a classic method to identify the causative genes of single-gene genetic diseases, but it also faces some difficulties: ① usually requires multi-generation families, and it is difficult to analyze small families and sporadic cases
②Sometimes multi-generational families cannot locate the pathogenic locus
③ It is difficult to screen out the correct disease-causing gene in the linkage region

Method used

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  • Method for screening HRDs disease-causing mutation and gene chip hybridization probe designing method involved in same
  • Method for screening HRDs disease-causing mutation and gene chip hybridization probe designing method involved in same
  • Method for screening HRDs disease-causing mutation and gene chip hybridization probe designing method involved in same

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] experimental method:

[0049] 1. Establishment of HRDs genetic resource bank.

[0050] 1.1 Collect the clinical data and blood samples of the following three types of patients:

[0051] 1.1.13 or more generations of autosomal dominant, autosomal recessive, and X-linked recessive families, including RP, Leber congenital amaurosis, congenital static night blindness, and yolk-like macular dystrophy , Stargardt disease.

[0052] 1.1.2 Collect small hereditary families of various HRDs.

[0053] 1.1.3 Collect sporadic cases of various HRDs without family history.

[0054] 1.2 Genomic DNA extraction:

[0055] Using TIANamp Blood DNA Extraction Kit (Tiangen Biotech Co. Ltd, Beijing, China), the patient's genomic DNA was extracted from the collected patient's peripheral blood according to the protocol provided by the manufacturer.

[0056] 2. Mining new pathogenic genes / new mutations of known pathogenic genes in HRDs (see figure 2 ).

[0057] 2.1 Design and customize HRDs...

Embodiment 2

[0096] Functional research was carried out on the pathogenic gene detected in Example 1. Here, the new mutation p.P315L of the above-mentioned PRPF4 gene was taken as an example.

[0097] experimental method:

[0098] 1. Conservative analysis:

[0099] Using NCBI HomoloGene database ( http: / / www.ncbi.nlm.nih.gov / homologene ) Conservative prediction of the screened mutations in multiple species.

[0100] 2. Predict the pathogenicity of mutations based on SIFT and PolyPhen values:

[0101] Using two mainstream online prediction software: PolyPhen-2 (Polymorphism Phenotyping, version2;

[0102] http: / / genetics.bwh.harvard.edu / pph2 / ) and SIFT Human Protein DB ( http: / / sift.bii.a-star.edu.sg / ), to predict the impact of missense mutations and nonsense mutations on protein levels, thereby predicting the pathogenicity of mutations.

[0103] 3. Research on protein crystal structure changes:

[0104] Since the PRPF4 gene is involved in the formation of the U4 / U6 complex, and...

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Abstract

The invention belongs to the field of biological medicines, and relates to a method for screening HRDs disease-causing mutation and a gene chip hybridization probe designing method involved in the same. The method for screening the HRDs disease-causing mutation comprises the steps of (1) establishing an HRDs genetic resource repository; (2) designing and synthesizing a gene chip hybridization probe of an HRDs disease-causing gene, and integrating the gene chip hybridization probe onto a gene chip; (3) capturing a target area by utilizing the prepared gene chip and executing the depth sequencing; (4) analyzing the sequencing data on the aspect of bioinformatics, and screening the candidate disease-causing gene; (5) functionally predicting a newly-discovered splicing gene mutation site. By establishing the high-efficient HRDs target gene capturing technology, adopting the depth sequencing as a means and confirming the efficiency of the HRDs capturing chip, a high-efficient credible biological information analysis model is established.

Description

technical field [0001] The invention belongs to the field of biomedicine and relates to a method for screening pathogenic mutations of HRDs and a method for designing hybridization probes for gene chips. Background technique [0002] Hereditary retinal diseases (HRDs) are a group of progressive retinal degenerative diseases caused by genetic defects, and are common and serious hereditary blinding diseases in clinical practice. As the number one blindness disease among working-age people worldwide, the incidence of HRDs is about 1 / 3000 in Europe and the United States, and as high as 1 / 1000 in my country. my country is a country with a large number of HRDs genetic resources, but most of the genetic information related to HRDs comes from western countries. Therefore, it is particularly important to conduct in-depth genetic research on HRDs patients in my country. [0003] HRDs are mostly single-gene genetic diseases, and many gene defects can lead to their occurrence. The comm...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11C12N15/10
CPCC12Q1/6811C12Q1/6869C12Q1/6883C12Q2600/156C12Q2565/519C12Q2535/122
Inventor 赵晨陈雪赵堪兴陈雪娟潘鑫源蒋超
Owner 赵晨
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