New mutant pathogenic gene of febrile convulsion as well as coding protein and application thereof

A gene and protein technology, applied in the field of new mutation-causing genes for febrile convulsions, can solve the problems of increased complexity, scalability, and complexity of the disease

Inactive Publication Date: 2015-02-25
BGI GENOMICS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] FS is genetically scalable and complex, resulting in increased complexity of the disease

Method used

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  • New mutant pathogenic gene of febrile convulsion as well as coding protein and application thereof
  • New mutant pathogenic gene of febrile convulsion as well as coding protein and application thereof
  • New mutant pathogenic gene of febrile convulsion as well as coding protein and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1: Screening of FS-related pathogenic gene loci from families with dominant inheritance of FS

[0043] 1. Sample collection

[0044] In recent years, the inventor of the present application has collected a four-generation Chinese family of FS dominant inheritance (such as figure 1 7 samples in ), including 3 patients (II-1, II-2, and Ⅲ-3) and 4 control samples (II-3, Ⅲ-4, Ⅳ-1 and Ⅳ-2). The disease diagnosis is based on fever The family history of sexual convulsions, clinical and biochemical indicators, and imaging evidence, combined with the familial cases collected by the inventor, suggest that FS is more likely. Take these 7 samples as research samples, and collect 2ml of peripheral blood samples for each sample, add EDTA for anticoagulation, and store at -80 degrees Celsius.

[0045] 2. DNA extraction

[0046] Use OMEGA Blood DNA Midi Kit whole blood DNA extraction kit to extract DNA from peripheral blood samples. The extraction steps are as follows:

[0047] (1) Tak...

Embodiment 2

[0075] Example 2: Sanger method sequencing verification

[0076] Sanger sequencing was used to verify the mutations detected by whole-exome sequencing on the PRRT2 gene in Example 1, and to verify the correlation between this mutant gene and FS disease. The PCR primers used to amplify the region of the mutation were Primer3 (http: / / frodo. wi.mit.edu / primer3 / ) design. The amplified fragments were sequenced using ABI3100 (Applied Biosystems, Foster City, CA) genetic analyzer and ABI BigDye Terminator cycle sequencing kit v3.1 (Applied Biosystems, Foster City, CA). The specific method steps are as follows:

[0077] ① DNA extraction

[0078] Take the peripheral venous blood of these 7 samples to extract genomic DNA according to the method in Example 1, and measure the DNA content spectrophotometrically.

[0079] ②Primer design and PCR reaction

[0080] First, refer to the human genome sequence database GRCh37.1 / hg19, and use Primer3.0 to design the exon-specific primers of PRRT2 gene res...

Embodiment 3

[0100] Example 3: In vitro detection of PRRT2 gene kit for FS patients

[0101] In order to detect pathogenic mutations in FS patients, all exons in the coding region of PRRT2 gene and primer sequences at the junction of exons and introns can be designed. In this example, the kit primers and amplification and sequencing conditions are as in Example 2.

[0102] 1. The composition of the kit:

[0103] Primer: as shown in Example 2;

[0104] Taq enzyme

[0105] Buffer

[0106] dNTP

[0107] Extract and purify genetic material (DNA samples) from biological samples to be tested

[0108] 2. How to use:

[0109] (1) Genomic DNA extraction: Use Wizard Genomic DNA extraction kit from Promega of the United States to extract genomic DNA from peripheral blood samples.

[0110] (2) First use the above PCR primers, Taq enzyme, sample genomic DNA, buffer, dNTP, etc. to perform PCR amplification reaction;

[0111] (3) Purify PCR amplification products;

[0112] (4) Perform BigDye reaction on the purified PCR...

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Abstract

The invention authenticates a new pathogenic gene-PRRT2 related with febrile convulsion. Specifically, a four-generation Chinese family of febrile convulsion dominant inheritance is used as a research object, sick individuals and non-sick individuals in the family are subjected to exome sequencing and comparison, a frame-shift insertion mutant c.641insC(p.R217Pfs*8) is discovered in the PRRT2 gene, and the mutant causes the change of a PRRT2 protein. Based on condition, the invention provides a mutant PRRT2 gene as well as a coding protein and application thereof, including a vector of the mutant PRRT2 gene, a host cell and a kit. The febrile convulsion can be subjected to molecular diagnosis and sickening risk evaluation by using the mutant PRRT2 gene. The mutant gene and the coding protein thereof can also be used as medicament targets for treating febrile convulsion.

Description

Technical field [0001] The invention relates to a human variant gene, in particular to a new mutant pathogenic gene for febrile convulsions. The present invention also relates to the coding protein and application of the new mutant pathogenic gene of febrile seizures, and the vector, host cell and kit containing the mutant febrile seizure pathogenic gene. Background technique [0002] Convulsions are commonly called cramps, convulsions, convulsions, or convulsions. Mainly manifested as paroxysmal twitching of the limbs and facial muscles, often accompanied by upturned eyeballs, staring or strabismus on both sides, and unconsciousness. Sometimes it is accompanied by foaming at the mouth or movement of the corners of the mouth, apnea, and purple complexion. The attack time is usually within 3 to 5 minutes. It is a common emergency in children, especially in infants and young children. Febrile seizures (FS) refers to infants and young children who have increased body temperature ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/12C07K14/47C12Q1/68C07K16/18A61K39/395A61P25/08
Inventor 管李萍肖晶晶谌于蓝邓昊徐洪波宋治郑文
Owner BGI GENOMICS CO LTD
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