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Gene chip for screening primary cilium-related disease causative gene

A gene chip and pathogenic gene technology, which is applied in the field of gene chip for the screening of pathogenic genes of primary cilia-related diseases, can solve the problems of diagnosis dependence, phenotype diagnosis interference, etc.

Inactive Publication Date: 2016-01-20
GENERAL HOSPITAL OF PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there is no reliable and definite treatment for primary cilia disease, and its diagnosis still depends on the clinical phenotype. Considering the complexity of the clinical phenotype, the pure phenotype diagnosis will bring some interference

Method used

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  • Gene chip for screening primary cilium-related disease causative gene
  • Gene chip for screening primary cilium-related disease causative gene
  • Gene chip for screening primary cilium-related disease causative gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Preparation of Example 1 Gene Chip and Preservation of Probes

[0032] 1. Probe synthesis

[0033] Probes of corresponding lengths were designed and synthesized by using in situ synthesis technology according to the probe sequences. The probe sequences are shown in SEQ ID NO.1-113, and the synthesized probes were purified by HPLC technology.

[0034] 2. Elution probe

[0035] Soak the chip with the probe in 5ml of 35% ammonia water, seal it and place it on a constant temperature rotator, rotate for more than 2 hours, elute the probe from the chip, and transfer it to centrifuge tube 2.0.

[0036] 3. Centrifugal drying

[0037] The eluted probe was concentrated in a rotary vacuum using a vacuum concentrator, and stopped when there was no liquid.

[0038] 4. Dissolution of the probe

[0039] Add 200 μl of RNA and DNase-free deionized water to the concentrated probe tube, vortex for more than 1 minute, and combine all dissolved probes into one tube for storage.

[0040...

Embodiment 2

[0044] The usage method of embodiment 2 gene chip

[0045] (1) Preparation of sample DNA fragments

[0046] The blood of patients and normal people was taken, and the whole genome DNA (gDNA) was extracted by conventional methods.

[0047] Adjust the temperature of the water bath to 37°C and 65°C, dissolve 10× buffer C (see the table below), vortex briefly and then centrifuge for a few seconds.

[0048] For each reaction, gDNA was added to an appropriate nuclease-free PCR tube and nuclease-free water was added to bring the reaction volume to 10.1 μl.

[0049] Prepare the enzyme digestion mix master solution on ice, and add the reagents in the order listed in Table 1:

[0050] Table 1

[0051] Reagent

Each reaction (μl)

×16rxns(μl)

water

1.0

17.0

[0052] 10× Buffer C

1.3

22.1

Acetylated BSA (10μg / μl)

0.1

1.7

Alu I (10U / μl)

0.25

4.25

Rsa I (10U / μl)

0.25

4.25

final vol...

Embodiment 3

[0081] Example 3 Detection Accuracy of Gene Chips to Primary Ciliary-Related Disease-causing Genes

[0082] 1. Selection of experimental subjects: 50 cases with clinical manifestations of primary cilia-related diseases were selected as test subjects, and the causative genes of the above test subjects have been determined using gene expression profiling and gene sequencing techniques.

[0083] 2. Utilize the gene chip of the present invention to detect the pathogenic gene of the case

[0084] According to the method described in Example 2, the pathogenic genes of the above cases were determined. The results show that: the gene chip containing the probe sequence shown in SEQIDNO.1-113 detected the disease-causing genes of 50 cases of patients with the same detection results obtained by using gene expression profile and gene sequencing technology, the gene chip is related to the primary cilia The detection accuracy of disease-causing genes is 100%.

[0085] 3. Comparative exper...

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Abstract

The invention discloses a gene chip for screening a primary cilium-related disease causative gene. The gene chip comprises hundreds of probes. The probes are used for detecting a primary cilium-related disease related gene. The gene chip realizes purposeful gene screening on primary cilium-related disease clinical phenotype cases, can fast and effective identify causative genes, and provides basis for primary cilium-related disease early diagnosis and individual treatment.

Description

technical field [0001] The invention belongs to the field of gene diagnosis and relates to a gene chip, in particular to a gene chip for primary cilia-related disease-causing gene screening and its application. Background technique [0002] Primary cilia-associated disorders are a group of overlapping clinical phenotypes and genotypes that are caused by defects in primary cilia. Cilia-related diseases involve various organs of the human body such as kidneys, brain, limbs, eyes, ears, liver, bones, etc. Its phenotypes include polycystic kidney disease, hepatobiliary disease, polydactyly, cognitive impairment, retinal degeneration, skeletal abnormalities, obesity, and visceral transposition. Primary cilia-related disorders include: Bardet-Biedl syndrome (BBS), juvenile wasting nephropathy (NPHP), Senior-Loken syndrome (SLNS), Alstrom syndrome (ALMS), Meckel-Grubber syndrome (MKS), Joubert syndrome Syndrome (JBTS), Orofacial-Digital Syndrome Type 1 (OFD1), Elliott Syndrome (E...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C40B40/06C12Q1/68
Inventor 卢彦平
Owner GENERAL HOSPITAL OF PLA
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